@article{miller_2021, title={Cultural Entanglements: Langston Hughes and the Rise of African and Caribbean Literature}, volume={118}, ISSN={["1545-6951"]}, DOI={10.1086/711904}, abstractNote={Previous article FreeBook ReviewCultural Entanglements: Langston Hughes and the Rise of African and Caribbean Literature. Shane Graham. Charlottesville: University of Virginia Press, 2020. Pp. vii+307.W. Jason MillerW. Jason MillerNC State University Search for more articles by this author Full TextPDF Add to favoritesDownload CitationTrack CitationsPermissionsReprints Share onFacebookTwitterLinked InRedditEmailQR Code SectionsMoreCultural Entanglements leads readers to ask, “What role should Langston Hughes play in postcolonial literature?” This book, the twelfth installment in the New World Studies series, turns on the reliable fact that, in 1943, Hughes started looking overseas to Africa rather than to a USSR that had preoccupied his attentiveness throughout the previous decade. Lest Africa seem like an obvious attraction for any black writer from the United States, we note that neither James Baldwin nor Richard Wright showed any interest in that continent, and Hughes was in fact the only writer of the New Negro Movement to walk its shores. Hughes’s trilinguarity as a speaker of French, Spanish, and English made him more equipped than most for this task. As a result, the author maps Hughes’s material connections across the African diaspora into the Caribbean (and beyond) to trace how Hughes very specifically became entangled with Claude McKay, Jacques Roumain, Aimé Césaire, Peter Abrahams, Es’kia Mphahlele, and Paule Marshall. Even the most ardent scholar will be pleasantly surprised at the staggering reciprocity Hughes experienced with the world outlined here (including France’s Négritude movement). Most welcomed is the ability to track Hughes’s reach beyond his death in 1967 so that writers of the late 1970s and ’80s are considered with new zeal. This tact culminates with the final study documenting the impact Hughes had on Paule Marshall’s 2009 memoir Triangular Road.An additional note of praise is that Graham astutely makes use of author interviews just as much as published works, so that direct nods to Hughes are captured in the first-person voice of the actual author (as opposed to more oblique references via fictional narrators). As expected, we find here all the names that sound the loudest in the fields of transatlantic and postcolonial studies.Cultural Entanglements excels when the author interrogates the inherited translations we have received. Variations are frequently explored with authority and balance, so that readers are left all the better for understanding what the French term “catholique” most likely suggested in Césasire’s original 1939 edition of his Notebook (134).Haiti reappears in several creative projects, often falling on a scale between primitivism and personhood. For many, the Haitian Revolution of 1791 (and its aftermath) serve as narratives to repeal modern states of oppression and hegemony. In the end, Haiti has often been portrayed with such vigor because it is seen as a failed opportunity to establish a truly native state of nationhood.The author clearly has his hands near the pulse of what is alive and happening in the worlds of modernist studies, Langston Hughes, and postcolonial literature as he has selected Hughes’s timely play/opera Emperor of Haiti (1936) / Troubled Island (1949) and his still-underexamined poetic sequence Ask Your Mama (1961) for in-depth exploration. Furthermore, the insightful interplay here between Césaire and Frantz Fanon reminds us that the student in this relationship continues to become more and more seminal to varying studies with each passing year.As to style, the text is often self-referential, pointing readers to its other sections (as if the text were expectedly prepared to be read electronically as single chapters). Readers will also have to wade through an inordinate number of early tropes that play off the idea of yarn and thread. This is the most noticeable when terms such as “arteries” sometimes appear twice in the same sentence (39–40), as do “knotted” and “knot” (50). In another instance, a “ship” is somehow both a “ship” and a “channel” in the same breath (43). These reoccurrences may say more about this field of literary studies than they do about the author. Fortunately, the redundancy wanes substantially as more exciting opportunities arise when Roumain’s powerful notion in “Ebony Wood” (1931) of a memory holding “Like a splinter in the wood” fixes as an emblem of the abject to extend the central framework of entanglement (and conversation) into the realms of contamination (92).Cultural Entanglements traces how so many of these authors worked to be detribalized and denationalized. While least interesting as applied to Claude McKay, this discussion is at its best when Césasire and Marshall are held in Graham’s nets. Even more impressive is the coalescence in chapter 4 of Abrahams, Mphahlele, Richard Rive, Hughes, and Drum magazine (for which Hughes served as literary contest judge in 1953). This is territory Graham knows better than anyone, and these sections are welcome additions to his earlier contribution as coeditor of Langston Hughes and the South African “Drum” Generation: The Correspondence (2010). While it could be stated more explicitly that Hughes’s political status in the United States gave him additional impetus to look across the Atlantic, we nonetheless realize that Hughes’s work as editor and judge was every bit as important to our reassessment of him as his need to dock overtly political poems such as “Not for Publication” (1959) overseas in outlets such as Nigeria’s Black Orpheus.Graham has the authority to assert that Hughes’s edited An African Treasury (1960) usurped an earlier edition of works by Africans because only Hughes had enough interaction with the continent to collect contemporary stories rather than old oral tales and legends. And these entanglements create fun everywhere as Mphahlele writes back to Hughes that the American has captured the spirit of the continent as “a giant rubbing his eyes as he walks,” so that astute readers note that the compliment is actually turning Hughes’s own imagery from his poem “Africa” (1952) right back to its author’s eyes (169).Cultural Entanglements reminds us that Hughes knew both Kwame Nkrumah and Nnamdi Azikiwe. The poet even attended the latter’s inauguration ceremonies with Martin Luther King Jr., where Hughes’s poem “Youth” served as the final section of the governor-general’s acceptance speech in 1960. Nonetheless, the poet still misspelled the name of Azikiwe’s son in Ask Your Mama, writing “AMEKA” instead of “Emeka” (206). Even without reference to this sequence’s first critic, Jean Wagner, Graham offers remarkable insight into “Gospel Cha-Cha” by offering a detailed contextual reading that draws upon a deep knowledge of Vodou and Afro-Jamaican religious sects popular throughout the Caribbean. Given these astute referents, the text is illuminated with a much broader sense of what Ask Your Mama is pointing to when it repeats “in the quarter of the Negroes” in twin dactylic feet (202). Along the way, Graham casually directs us to an unpublished poem Hughes wrote about Roumain that immediately sparks our interest to explore it in even greater detail.In short, Cultural Entanglements is a primer for where things are currently developing between Hughes and our understanding of postcolonial literature. It is also a ready-made resource waiting to be plumbed by scholars now forming new studies into the ever-prescient Hughes and his vast global network. Previous article DetailsFiguresReferencesCited by Modern Philology Volume 118, Number 3February 2021 Article DOIhttps://doi.org/10.1086/711904 Views: 287 HistoryPublished online October 30, 2020 For permission to reuse, please contact [email protected] Crossref reports no articles citing this article.}, number={3}, journal={MODERN PHILOLOGY}, author={Miller, W. Jason}, year={2021}, month={Feb}, pages={E224–E226} }
 @article{fan_foster_miller_osborne_kathariou_2021, title={Impact of Ceftiofur Administration in Steers on the Prevalence and Antimicrobial Resistance of Campylobacter spp.}, volume={9}, ISSN={["2076-2607"]}, DOI={10.3390/microorganisms9020318}, abstractNote={Bacterial resistance to ceftiofur raises health concerns due to ceftiofur’s extensive veterinary usage and structural similarity with the human antibiotic ceftriaxone. Ceftiofur crystalline-free acid (CCFA) and ceftiofur hydrochloride (CHCL) are ceftiofur types used therapeutically in cattle, but their potential impacts on Campylobacter prevalence and antimicrobial resistance remain unclear. In this study two groups of steers were each treated with CCFA or CHCL. In vivo active drug concentrations were measured and fecal samples were analyzed for Campylobacter for up to 42 days post-treatment. Following administration, the colonic concentration of ceftiofur initially increased then dropped to pre-treatment levels by day 8. The estimated prevalence of Campylobacter spp. was significantly (p = 0.0009) higher during the first week after CCFA treatment than after CHCL treatment (81.3% vs. 45.2%). Campylobacter jejuni predominated overall, with other Campylobacter spp. mainly identified in the first week after CCFA treatment. No treatment impacts were noted on ceftiofur minimum inhibitory concentration (MIC) for C. jejuni (10–20 μg/mL). More C. jejuni genotypes were detected in CCFA-treated than CHCL-treated steers. These findings suggest that ceftiofur did not significantly impact Campylobacter prevalence or ceftiofur MIC. However, CHCL may be preferable due to the lower likelihood of temporary increases in Campylobacter prevalence.}, number={2}, journal={MICROORGANISMS}, author={Fan, Sicun and Foster, Derek and Miller, William G. and Osborne, Jason and Kathariou, Sophia}, year={2021}, month={Feb} }
 @article{good_miller_niedermeyer_osborne_siletzky_carver_kathariou_2019, title={Strain-Specific Differences in Survival of Campylobacter spp. in Naturally Contaminated Turkey Feces and Water}, volume={85}, ISSN={["1098-5336"]}, DOI={10.1128/AEM.01579-19}, abstractNote={Campylobacter jejuni and Campylobacter coli are leading foodborne pathogens, with poultry as a major reservoir. Due to their growth requirements, these Campylobacter spp. may be unable to replicate once excreted by their avian hosts, but their survival in feces and the environment is critical for transmission in the farm ecosystem. Reducing the prevalence of Campylobacter-positive flocks can have major impacts in controlling both contamination of poultry products and environmental dissemination of the pathogens. However, understanding the capacity of these pathogens to survive in transmission-relevant vehicles such as feces and farmhouse water remains poorly understood, and little information is available on species- and strain-associated differences in survival. Here, we employed model conditions to investigate the survival of C. jejuni and C. coli from naturally colonized turkey flocks, and with diverse genotypes and antimicrobial resistance profiles, in turkey feces and in farmhouse water. ABSTRACT Campylobacter jejuni and Campylobacter coli are leading causes of human foodborne illness, with poultry as a major vehicle. Turkeys are frequently colonized with Campylobacter, but little is known about Campylobacter survival in turkey feces, even though fecal droppings are major vehicles for Campylobacter within-flock transmission as well as for environmental dissemination. Our objective was to examine survival of Campylobacter, including different strains, in freshly excreted feces from naturally colonized commercial turkey flocks and in suspensions of turkey feces in water from the turkey house. Fecal and water suspensions were stored at 4°C, and Campylobacter populations were enumerated on selective media at 48-h intervals. C. jejuni and C. coli isolates were characterized for resistance to a panel of antibiotics, and a subset was subtyped using multilocus sequence typing. Campylobacter was recovered from feces and water for up to 16 days. Analysis of 548 isolates (218 C. jejuni and 330 C. coli) revealed that C. jejuni survived longer than C. coli in feces (P = 0.0005), while the reverse was observed in water (P < 0.0001). Strain-specific differences in survival were noted. Multidrug-resistant C. jejuni isolates of sequence type 1839 (ST-1839) and the related ST-2935 were among the longest-surviving isolates in feces, being recovered for up to 10 to 16 days, while multidrug-resistant C. coli isolates of ST-1101 were recovered from feces for only up to 4 days. Data on Campylobacter survival upon excretion from the birds can contribute to further understanding of the transmission dynamics of this pathogen in the poultry production ecosystem. IMPORTANCE Campylobacter jejuni and Campylobacter coli are leading foodborne pathogens, with poultry as a major reservoir. Due to their growth requirements, these Campylobacter spp. may be unable to replicate once excreted by their avian hosts, but their survival in feces and the environment is critical for transmission in the farm ecosystem. Reducing the prevalence of Campylobacter-positive flocks can have major impacts in controlling both contamination of poultry products and environmental dissemination of the pathogens. However, understanding the capacity of these pathogens to survive in transmission-relevant vehicles such as feces and farmhouse water remains poorly understood, and little information is available on species- and strain-associated differences in survival. Here, we employed model conditions to investigate the survival of C. jejuni and C. coli from naturally colonized turkey flocks, and with diverse genotypes and antimicrobial resistance profiles, in turkey feces and in farmhouse water.}, number={22}, journal={APPLIED AND ENVIRONMENTAL MICROBIOLOGY}, author={Good, Lesley and Miller, William G. and Niedermeyer, Jeffrey and Osborne, Jason and Siletzky, Robin M. and Carver, Donna and Kathariou, Sophia}, year={2019}, month={Nov} }
 @article{young_oh_williams_foster_miller_lunn_mowat_2018, title={Clinical therapeutic efficacy of mycophenolate mofetil in the treatment of SARDS in dogs-a prospective open-label pilot study}, volume={21}, ISSN={["1463-5224"]}, DOI={10.1111/vop.12545}, abstractNote={Abstract}, number={6}, journal={VETERINARY OPHTHALMOLOGY}, author={Young, Whitney M. and Oh, Annie and Williams, Jonathan G. and Foster, Melanie L. and Miller, William W. and Lunn, Katharine F. and Mowat, Freya M.}, year={2018}, month={Nov}, pages={565–576} }
 @article{niedermeyer_ring_miller_genger_lindsey_osborne_kathariou_2018, title={Proximity to Other Commercial Turkey Farms Affects Colonization Onset, Genotypes, and Antimicrobial Resistance Profiles of Campylobacter spp. in Turkeys: Suggestive Evidence from a Paired-Farm Model}, volume={84}, ISSN={["1098-5336"]}, DOI={10.1128/AEM.01212-18}, abstractNote={
            Colonization of poultry with
            Campylobacter
            at the farm level is complex, poorly understood, and critically linked to contamination of poultry products, which is known to constitute a leading risk factor for human campylobacteriosis. Here, we investigated the use of a paired-farm design under standard production conditions and in the absence of experimental inoculations to assess potential impacts of farm and host genetics on prevalence, antimicrobial resistance and genotypes of
            Campylobacter
            in commercial turkeys of two different breeds. Data suggest impacts of farm proximity to other commercial turkey farms on the onset of colonization, genotypes, and antimicrobial resistance profiles of
            Campylobacter
            colonizing the birds. Furthermore, the significant association of a specific multidrug-resistant
            Campylobacter jejuni
            strain with turkeys of one breed suggests colonization partnerships at the
            Campylobacter
            strain-turkey breed level. The study design avoids potential pitfalls associated with experimental inoculations, providing novel insights into the dynamics of turkey colonization with
            Campylobacter
            in actual farm ecosystems.
          }, number={18}, journal={APPLIED AND ENVIRONMENTAL MICROBIOLOGY}, author={Niedermeyer, Jeffrey A. and Ring, Lynde and Miller, William G. and Genger, Seiche and Lindsey, Christina Parr and Osborne, Jason and Kathariou, Sophia}, year={2018}, month={Sep} }
 @article{wang_hastings_miller_kumar_2016, title={Fshb-iCre mice are efficient and specific Cre deleters for the gonadotrope lineage}, volume={419}, ISSN={["0303-7207"]}, DOI={10.1016/j.mce.2015.10.006}, abstractNote={Genetic analysis of development and function of the gonadotrope cell lineage within mouse anterior pituitary has been greatly facilitated by at least three currently available Cre strains in which Cre was either knocked into the Gnrhr locus or expressed as a transgene from Cga and Lhb promoters. However, in each case there are some limitations including CRE expression in thyrotropes within pituitary or ectopic expression outside of pituitary, for example in some populations of neurons or gonads. Hence, these Cre strains often pose problems with regard to undesirable deletion of alleles in non-gonadotrope cells, fertility and germline transmission of mutant alleles. Here, we describe generation and characterization of a new Fshb-iCre deleter strain using 4.7 kb of ovine Fshb promoter regulatory sequences driving iCre expression exclusively in the gonadotrope lineage within anterior pituitary. Fshb-iCre mice develop normally, display no ectopic CRE expression in gonads and are fertile. When crossed onto a loxP recombination-mediated red to green color switch reporter mouse genetic background, in vivo CRE recombinase activity is detectable in gonadotropes at more than 95% efficiency and the GFP-tagged gonadotropes readily purified by fluorescence activated cell sorting. We demonstrate the applicability of this Fshb-iCre deleter strain in a mouse model in which Dicer is efficiently and selectively deleted in gonadotropes. We further show that loss of DICER-dependent miRNAs in gonadotropes leads to profound suppression of gonadotropins resulting in male and female infertility. Thus, Fshb-iCre mice serve as a new genetic tool to efficiently manipulate gonadotrope-specific gene expression in vivo.}, number={C}, journal={MOLECULAR AND CELLULAR ENDOCRINOLOGY}, author={Wang, Huizhen and Hastings, Richard and Miller, William L. and Kumar, T. Rajendra}, year={2016}, month={Jan}, pages={124–138} }
 @article{wang_larson_jablonka-shariff_pearl_miller_conn_boime_kumar_2014, title={Redirecting intracellular trafficking and the secretion pattern of FSH dramatically enhances ovarian function in mice}, volume={111}, ISSN={["0027-8424"]}, DOI={10.1073/pnas.1321404111}, abstractNote={Significance}, number={15}, journal={PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA}, author={Wang, Huizhen and Larson, Melissa and Jablonka-Shariff, Albina and Pearl, Christopher A. and Miller, William L. and Conn, P. Michael and Boime, Irving and Kumar, T. Rajendra}, year={2014}, month={Apr}, pages={5735–5740} }
 @article{jia_shafiee-kermani_miller_2013, title={Gonadotrope-Specific Expression and Regulation of Ovine Follicle Stimulating Hormone Beta: Transgenic and Adenoviral Approaches Using Primary Murine Gonadotropes}, volume={8}, ISSN={["1932-6203"]}, DOI={10.1371/journal.pone.0066852}, abstractNote={The beta subunit of follicle stimulating hormone (FSHB) is expressed specifically in pituitary gonadotropes in vertebrates. Transgenic mouse studies have shown that enhancers in the proximal promoter between −172/−1 bp of the ovine FSHB gene are required for gonadotrope expression of ovine FSHB. These enhancers are associated with regulation by activins and gonadotropin releasing hormone (GnRH). Additional distal promoter sequence between −4741/−750 bp is also required for expression. New transgenic studies presented here focus on this distal region and narrow it to 1116 bp between −1866/−750 bp. In addition, adenoviral constructs were produced to identify these critical distal sequences using purified primary mouse gonadotropes as an in vitro model system. The adenoviral constructs contained −2871 bp, −750 bp or −232 bp of the ovine FSHB promoter. They all showed gonadotrope-specific regulation since they were induced only in purified primary gonadotropes by activin A (50 ng/ml) and inhibited by GnRH (100 nM) in the presence of activin (except −232FSHBLuc). However, basal expression of all three viral constructs (in the presence of follistatin to block cellular induction by activin) was relatively high in pituitary non-gonadotropes as well as gonadotropes. Thus, gonadotrope-specific regulation associated with the proximal promoter was observed as expected, but the model was blind to distal promoter elements between −2871/−750 necessary for gonadotrope-specific expression of ovine FSHB in vivo. The new adenoviral-based in vitro technique did detect, however, a novel GnRH response element between −750 bp and −232 bp of the ovine FSHB promoter. We conclude that adenoviral-based studies in primary gonadotropes can adequately recognize regulatory elements on the ovine FSHB promoter associated with gonadotrope-specific regulation/expression, but that more physiologically based techniques, such as transgenic studies, will be needed to identify sequences between −1866/−750 bp of the ovine FSHB promoter that are also required for tissue/cell specific expression in vivo.}, number={7}, journal={PLOS ONE}, author={Jia, Jingjing and Shafiee-Kermani, Farideh and Miller, William L.}, year={2013}, month={Jul} }
 @article{han_miller_2009, title={Activin A induces ovine follicle stimulating hormone beta using-169/-58 bp of its promoter and a simple TATA box}, volume={7}, ISSN={["1477-7827"]}, DOI={10.1186/1477-7827-7-66}, abstractNote={Activin A increases production of follicle stimulating hormone (FSH) by inducing transcription of its beta subunit (FSHB). This induction has been studied here in LbetaT2 gonadotropes using transient expression of ovine FSHBLuc (-4741 bp of ovine FSHB promoter plus exon/intron 1 linked to Luc). Several sequences between -169/-58 bp of the ovine FSHB proximal promoter are necessary for induction by activin A in LbetaT2 cells, but deletions between -4741/-752 bp decrease induction > 70% suggesting the existence of other important 5' sequences. Induction disappears if a minimal T81 thymidine kinase promoter replaces the ovine FSHB TATA box and 3' exon/intron. The study reported here was designed to determine if sequences outside -169/-58 bp are important for induction of ovine FSHB by activin A. Progressively longer deletions of ovine FSHBLuc were created between -4741/-195 bp. Deletions internal to this region were created also, but replaced with substitute DNA. The ovine FSHB TATA box region (-40/+3 bp) was replaced by thymidine kinase and rat prolactin minimal promoters, and substitutions were made in 3' intron/exon sequences. All constructs were tested for basal and activin A-induced expression in LbetaT2 cells. Successive 5' deletions progressively lowered fold-induction by activin A from 9.5 to zero, but progressively increased basal expression. Replacing deletions with substitute DNA showed no changes in basal expression or fold-induction. Induction by activin A was supported by the minimal rat prolactin promoter (TATA box) but not the thymidine kinase promoter (no TATA box). Replacement mutations in the 3' region did not decrease induction by activin A. The data show that specific ovine FSHB sequences 5' to -175 bp or 3' of the transcription start site are not required for induction by activin A. A minimal TATA box promoter supports induction by activin A, but the sequence between the TATA box and transcription start site seems unimportant.}, journal={REPRODUCTIVE BIOLOGY AND ENDOCRINOLOGY}, author={Han, Sang-Oh and Miller, William L.}, year={2009}, month={Jun} }
 @misc{miller_2009, title={Increasing gamete production with a gene switch}, volume={7,579,515}, number={2009 Aug. 25}, author={Miller, W. L.}, year={2009} }
 @article{shafiee-kermani_han_miller_2007, title={Chronic gonadotropin-releasing hormone inhibits activin induction of the ovine follicle-stimulating hormone beta-subunit: Involvement of 3 ',5 '-cyclic adenosine monophosphate response element binding protein and nitric oxide synthase type I}, volume={148}, ISSN={["1945-7170"]}, DOI={10.1210/en.2006-1740}, abstractNote={FSH is induced by activin, and this expression is modulated by GnRH through FSHB expression. This report focuses on the inhibitory effect of GnRH on activin-induced FSHB expression. Activin-treated primary murine pituitary cultures robustly express mutant ovine FSHBLuc-ΔAP1, a luciferase transgene driven by 4.7 kb of ovine FSHB promoter. This promoter lacks two GnRH-inducible activator protein-1 sites, making it easier to observe GnRH-mediated inhibition. Luciferase expression from this transgene was decreased 94% by 100 nm GnRH with a half-time of approximately 4 h in pituitary cultures, and this inhibition was independent of follistatin. Activators of cAMP and protein kinase C like forskolin and phorbol 12-myristate 3-acetate (PMA), respectively, mimicked GnRH action. Kinetic studies of wild-type ovine FSHBLuc in LβT2 cells showed continuous induction by activin (4-fold) over 20 h. Most of this induction (78%) was blocked, beginning at 6 h. cAMP response element binding protein (CREB) was implicated in this inhibition because overexpression of its constitutively active mutant mimicked GnRH, and its inhibitor (inducible cAMP early repressor isoform II) reversed the inhibition caused by GnRH, forskolin, or PMA. In addition, GnRH, forskolin, or PMA increased the expression of a CREB-responsive reporter gene, 6xCRE-37PRL-Luc. Inhibition of nitric oxide type I (NOSI) by 7-nitroindazole also reversed GnRH-mediated inhibition by 60%. It is known that GnRH and CREB induce production of NOSI in gonadotropes and neuronal cells, respectively. These data support the concept that chronic GnRH inhibits activin-induced ovine FSHB expression by sequential activation of CREB and NOSI through the cAMP and/or protein kinase C pathways.}, number={7}, journal={ENDOCRINOLOGY}, author={Shafiee-Kermani, Farideh and Han, Sang-oh and Miller, William L.}, year={2007}, month={Jul}, pages={3346–3355} }
 @article{su_shafiee-kermani_gore_jia_wu_miller_2007, title={Expression and regulation of the beta-subunit of ovine follicle-stimulating hormone relies heavily on a promoter sequence likely to bind smad-associated proteins}, volume={148}, ISSN={["1945-7170"]}, DOI={10.1210/en.2006-1635}, abstractNote={FSH is essential for normal gonadal function in mammals. Expression of its beta-subunit (FSHB) controls overall production/secretion of FSH and is induced by activin. Studies with ovine FSHB promoter/reporter constructs in L beta T2 gonadotropes show that induction by activin requires a putative Smad binding element in the ovine FSHB promoter (-162AGAC-159). Similar studies reported here show that another site, juxtaposed to the Smad binding element, was also required for 81% activin induction in L beta T2 cells. This site was similar to several that bind proteins known to partner with Smads. When this site (-171ACTgcgtTT-163) was mutated by changing the nucleotides shown in lowercase letters, the resulting ovine-derived construct (mut-oFSHBLuc) was expressed poorly as a transgene in primary mouse gonadotropes (<0.001 times compared with ovine wild-type transgenes). This decrease in expression demonstrated the importance of this site for activin induction and, perhaps, basal expression, although studies with L beta T2 cells did not suggest this latter possibility. Expression of mut-oFSHBLuc in male mouse gonadotropes in vivo was at least 644 times greater than expression in all but one nongonadotrope tissue tested, indicating that mut-oFSHBLuc retained significant gonadotrope-specific expression. An increase in FSHB expression occurs during estrus in mice and is faithfully reproduced with wild-type ovine FSHBLuc transgenes, but not with mut-oFSHBLuc, indicating that the mutated site is needed for this secondary FSH surge. These data suggest that activin gathers Smads and Smad-associated proteins at the -171/-159 promoter region to regulate expression of the ovine FSHB and overall FSH production.}, number={9}, journal={ENDOCRINOLOGY}, author={Su, Pei and Shafiee-Kermani, Farideh and Gore, A. Jesse and Jia, Jingjing and Wu, Joyce C. and Miller, William L.}, year={2007}, month={Sep}, pages={4500–4508} }
 @article{su_wu_sommer_gore_petters_miller_2005, title={Conditional induction of ovulation in mice}, volume={73}, ISSN={["1529-7268"]}, DOI={10.1095/biolreprod.104.039164}, abstractNote={Abstract Follicle-stimulating hormone controls the maturation of mammalian ovarian follicles. In excess, it can increase ovulation (egg production). Reported here is a transgenic doxycycline-activated switch, tested in mice, that produced more FSHB subunit (therefore more FSH) and increased ovulation by the simple feeding of doxycycline (Dox). The transgenic switch was expressed selectively in pituitary gonadotropes and was designed to enhance normal expression of FSH when exposed to Dox, but to be regulated by all the hormones that normally control FSH production in vivo. Feeding maximally effective levels of Dox increased overall mRNA for FSHB and serum FSH by over half in males, and Dox treatment more than doubled the normal ovulation rate of female mice for up to 10 reproductive cycles. Lower levels of Dox increased the number of developing embryos by 30%. Ovarian structure and function appeared normal. In summary, gene switch technology and normal FSH regulation were combined to effectively enhance ovulation in mice. Theoretically, the same strategy can be used with any genetic switch to increase ovulation (or any highly conserved physiology) in any mammal.}, number={4}, journal={BIOLOGY OF REPRODUCTION}, author={Su, P and Wu, JC and Sommer, JR and Gore, AJ and Petters, RM and Miller, WL}, year={2005}, month={Oct}, pages={681–687} }
 @article{gore_philips_miller_bernard_2005, title={Differential regulation of follicle stimulating hormone by activin A and TGFB1 in murine gonadotropes}, volume={3}, journal={Reproductive Biology and Endocrinology}, author={Gore, A. J. and Philips, D. P. and Miller, W. L. and Bernard, D. J.}, year={2005} }
 @article{safwat_ninomiya-tsuji_gore_miller_2005, title={Transforming growth factor beta-activated kinase 1 is a key mediator of ovine follicle-stimulating hormone beta-subunit expression}, volume={146}, ISSN={["1945-7170"]}, DOI={10.1210/en.2005-0457}, abstractNote={FSH, a key regulator of gonadal function, contains a β-subunit (FSHβ) that is transcriptionally induced by activin, a member of the TGFβ-superfamily. This study used 4.7 kb of the ovine FSHβ-promoter linked to luciferase (oFSHβLuc) plus a well-characterized activin-responsive construct, p3TPLuc, to investigate the hypothesis that Smad3, TGFβ-activated kinase 1 (TAK1), or both cause activin-mediated induction of FSH. Overexpression of either Smad3 or TAK1 induced oFSHβLuc in gonadotrope-derived LβT2 cells as much as activin itself. Induction of p3TPLuc by activin is known to require Smad3 activation in many cell types, and this was true in LβT2 cells, where 10-fold induction by activin (2–8 h after activin treatment) was blocked more than 90% by two dominant negative (DN) inhibitors of Smad3 [DN-Smad3 (3SA) and DN-Smad3 (D407E)]. By contrast, 6.5-fold induction of oFSHβLuc by activin (10–24 h after activin treatment) was not blocked by either DN-Smad inhibitor, suggesting that activation of Smad3 did not trigger induction of oFSHβLuc. By contrast, inhibition of TAK1 by a DN-TAK1 construct led to a 50% decrease in activin-mediated induction of oFSHβLuc, and a specific inhibitor of TAK1 (5Z-7-Oxozeanol) blocked induction by 100%, indicating that TAK1 is necessary for activin induction of oFSHβLuc. Finally, inhibiting p38-MAPK (often activated by TAK1) blocked induction of oFSHβLuc by 60%. In conclusion, the data presented here indicate that activation of TAK1 (and probably p38-MAPK), but not Smad3, is necessary for triggering induction of oFSHβ by activin.}, number={11}, journal={ENDOCRINOLOGY}, author={Safwat, N and Ninomiya-Tsuji, J and Gore, AJ and Miller, WL}, year={2005}, month={Nov}, pages={4814–4824} }
 @article{wu_su_safwat_sebastian_miller_2004, title={Rapid, efficient isolation of murine gonadotropes and their use in revealing control of follicle-stimulating hormone by paracrine pituitary factors}, volume={145}, ISSN={["1945-7170"]}, DOI={10.1210/en.2004-0257}, abstractNote={FSH and LH are produced only in gonadotropes, which are reported to comprise 3-12% of mammalian pituitaries. Factors made within the pituitary are powerful regulators of FSH and also influence LH expression, but their identities and cellular origins are unknown because it is impossible to isolate and individually analyze different pituitary cell types. In this study FSH-producing gonadotropes were specifically tagged in vivo with a transgenic cell surface antigen (H-2Kk) so they could be purified in vitro using paramagnetic anti-H-2Kk microbeads. After enzymatic dispersion of pituitary cells, it took 1 h or less to extract 55 +/- 5% of FSH-producing gonadotropes at 95 +/- 0.5% purity, as judged by immunostaining for FSH or prolactin. Although this procedure selected for FSH expression, the isolated gonadotropes were also enriched 22-fold for LH-containing cells. For studies aimed at understanding factors that control FSH transcription, the purified gonadotropes were treated with activin A, which increased FSH expression 480% above basal levels (d 3 of culture). Coincubation of purified gonadotropes with pituitary nongonadotropes increased FSH expression 800% (d 3 of culture). Follistatin, an activin-binding protein, decreased FSH expression 35-50%, suggesting that gonadotropes make some activin and/or other follistatin-sensitive molecule(s) that induce FSH. These data show that paracrine factors from pituitary nongonadotropes can play a major role in controlling FSHbeta at the pituitary level. The study presented here describes a rapid, reliable, and efficient method for isolating any specialized cell type, including all cells that produce endocrine hormones.}, number={12}, journal={ENDOCRINOLOGY}, author={Wu, JC and Su, P and Safwat, NW and Sebastian, J and Miller, WL}, year={2004}, month={Dec}, pages={5832–5839} }
 @article{pernasetti_spady_hall_rosenberg_givens_anderson_paulus_miller_mellon_2003, title={Pituitary tumorigenesis targeted by the ovine follicle-stimulating hormone beta-subunit gene regulatory region in transgenic mice}, volume={203}, ISSN={["0303-7207"]}, DOI={10.1016/S0303-7207(02)00430-6}, abstractNote={Targeted tumorigenesis in transgenic mice has been a powerful tool for the study of gene expression and oncogenesis, as well as for the production of differentiated immortal cell lines from rare cell types. Follicle-stimulating hormone (FSH) is secreted by the gonadotrope cells of the anterior pituitary gland and plays a pivotal role in mammalian reproduction. Here we have used the regulatory region of the ovine FSH beta gene to direct expression of the SV40 T antigen oncogene to gonadotrope cells in the pituitary of transgenic mice. Two of five transgenic mouse lines bearing this fusion gene rapidly developed pituitary tumors, with appearance of adenomatous foci as early as 6 weeks of age, resulting in death by 12 weeks of age in both genders. Histologic examination of tumor development over time revealed that increases in cell proliferation and dysplasia were accompanied by decreases in synthesis of pituitary hormones, indicating dedifferentiation of the pituitary cells. Histological features observed in these tumors were in agreement with this rapid transformation of cell phenotype. Tumors were multifocal in origin, and the most highly transformed cell types observed consisted of giant pale basophilic cells with enormous hyperploid nuclei associated with infiltrating neuronal-like cells, which were very abundant at later stages of tumor development. Mitotic indices were much higher in transgenic than wild-type pituitaries, as expected. Morphologic analysis of the gonads of these transgenic mice showed no major developmental differences, as compared to wild-type littermates, however the length of the seminiferous tubules in transgenic males was greater than age-matched wild-type animals. Despite this phenotype difference, both genders were fertile, with normal sperm development observed in males and normal estrous cycle stages in females. Moreover, while 8 -- 10-week-old transgenic males had much lower blood levels of FSH than littermates, transgenic female FSH levels were the same as those of wild-type females. These animals offer a unique and potentially useful model of organ-specific tumorigenesis, where a multistage pathway of tumor development is evident, both histologically and temporally. Study of such models will advance our knowledge on the physiological and molecular mechanisms involved in gene expression as well as tumor formation.}, number={1-2}, journal={MOLECULAR AND CELLULAR ENDOCRINOLOGY}, author={Pernasetti, F and Spady, TJ and Hall, SB and Rosenberg, SB and Givens, ML and Anderson, S and Paulus, M and Miller, WL and Mellon, PL}, year={2003}, month={May}, pages={169–183} }
 @misc{miller_shafiee-kermani_strahl_huang_2002, title={The nature of FSH induction by GnRH}, volume={13}, ISSN={["1043-2760"]}, DOI={10.1016/S1043-2760(02)00614-8}, abstractNote={Follicle-stimulating hormone (FSH), a major regulator of mammalian gonadal function, is induced by gonadotropin-releasing hormone (GnRH), but it is unclear how much induction is direct or indirect and what relevance each has in vivo. Two advances now make it possible to address these issues, which are central to understanding FSH regulation. The first is the use of transformed L beta T2 gonadotropes to define key promoter sequences of FSHB (the gene encoding the FSH-beta subunit) that are needed for induction by GnRH and/or other factors; and the second is the ability to express FSHB promoter-reporter constructs in transgenic mouse gonadotropes to test the physiological relevance of promoter elements identified by using L beta T2 cells. Here, we summarize past studies on GnRH induction of FSH, and propose questions and approaches for the future.}, number={6}, journal={TRENDS IN ENDOCRINOLOGY AND METABOLISM}, author={Miller, WL and Shafiee-Kermani, F and Strahl, BD and Huang, HJ}, year={2002}, month={Aug}, pages={257–263} }
 @article{huang_wu_su_zhirnov_miller_2001, title={A novel role for bone morphogenetic proteins in the synthesis of follicle-stimulating hormone}, volume={142}, ISSN={["1945-7170"]}, DOI={10.1210/en.142.6.2275}, abstractNote={FSH is produced in pituitary gonadotropes as an α/β heterodimer, and synthesis of the β-subunit is the rate-limiting step in overall FSH production. Synthesis of FSHβ can be regulated by activin and inhibin, both of which are members of the transforming growth factor-β superfamily. Bone morphogenetic proteins (BMPs) also belong to the transforming growth factor-β family and are multifunctional growth factors involved in many aspects of tissue development and morphogenesis, including regulation of FSH action in the ovary. Here we report a novel function for BMP-7 and BMP-6 in regulating FSH synthesis in the pituitary. Using primary pituitary cell cultures derived from transgenic mice that carry the ovine FSHβ promoter linked to a luciferase reporter gene (oFSHβLuc), BMP-7 or BMP-6 was found to stimulate oFSHβLuc expression by 6-fold. Transient expression of the oFSHβLuc in a transformed gonadotrope cell line, LβT2, was induced 4-fold by BMP-7 or BMP-6 treatment. More importantly, BMP-7 and BMP-6 increased endogenous FSH secretion by 10- and 14-fold, respectively, from LβT2 cells, demonstrating for the first time that a functional signaling BMP system is present in gonadotropes. Two bioneutralizing antibodies to BMP-7, which cross-react with BMP-6, but not with activin A, decreased basal oFSHβLuc expression and FSH secretion from transgenic mouse pituitary cultures by 83–88% and 47–48%, respectively, suggesting an autocrine or paracrine role for BMP-7 or BMP-6 in FSH synthesis. Neither bioneutralizing antibody to activin A or activin B decreased basal oFSHβLuc expression or mouse FSH secretion significantly. Dose-dependent inhibition of FSH synthesis by anti-BMP7 was also observed in rat and sheep pituitary cultures. These results combined with the fact that the messenger RNAs for BMP-7 and BMP-6 were detected in mouse pituitaries and LβT2 cells indicate that BMP-7 and/or BMP-6 can function as FSH stimulators and may be significant physiological factors maintaining basal FSH expression in vivo.}, number={6}, journal={ENDOCRINOLOGY}, author={Huang, HJ and Wu, JC and Su, P and Zhirnov, O and Miller, WL}, year={2001}, month={Jun}, pages={2275–2283} }
 @article{pernasetti_vasilyev_rosenberg_bailey_huang_miller_mellon_2001, title={Cell-specific transcriptional regulation of follicle-stimulating hormone-beta by activin and gonadotropin-releasing hormone in the L beta T2 pituitary gonadotrope cell model}, volume={142}, ISSN={["0013-7227"]}, DOI={10.1210/en.142.6.2284}, abstractNote={FSH is secreted by gonadotropes of the anterior pituitary and plays a crucial role in mammalian reproduction. However, little is known about FSH gene regulation due to the lack of a gonadotrope cell line that synthesizes FSH. The LβT2 mouse pituitary cell line, isolated by targeted tumorigenesis in transgenic mice, has the characteristics of a mature gonadotrope, including expression of GnRH receptor, steroidogenic factor 1, and both the α- and β-subunits of LH, but was thought not to express FSH. Using RT-PCR, we show that these cells synthesize FSH β- subunit messenger RNA, which is induced by activin and inhibited by follistatin. Furthermore, in transient transfections an ovine FSHβ 5′-regulatory region (5.5 kb) confers LβT2 cell-specific expression to a reporter gene compared with other pituitary and nonpituitary cell lines. This FSHβ regulatory region responds to activin specifically in LβT2 cells, an effect that is blocked by follistatin. The LHβ, α-subunit, and GnRH receptor regulatory regions are induced by activin and blocked by follistatin. Furthermore, LβT2 cells express the components of the activin system, and addition of follistatin alone reduces FSHβ gene expression, demonstrating that an endogenous activin autocrine loop regulates FSH in these cells. In addition, GnRH stimulates both the FSHβ and LHβ regulatory regions, specifically in LβT2 cells. Surprisingly, GnRH induction is reduced by follistatin, suggesting its dependence on endogenous activin. As the mouse GnRH receptor promoter is inhibited by follistatin, reduction of GnRH receptor levels might be one mechanism by which follistatin interferes with GnRH induction of gonadotropin genes. In summary, LβT2 cells exhibit the characteristics of fully differentiated gonadotropes, including the expression of LH, FSH, GnRH receptor, and components of the activin/follistatin system, as well as display the appropriate responses to activin and GnRH.}, number={6}, journal={ENDOCRINOLOGY}, author={Pernasetti, F and Vasilyev, VV and Rosenberg, SB and Bailey, JS and Huang, HJ and Miller, WL and Mellon, PL}, year={2001}, month={Jun}, pages={2284–2295} }
 @article{huang_sebastian_strahl_wu_miller_2001, title={The promoter for the ovine follicle-stimulating hormone-beta gene (FSH beta) confers FSH beta-like expression on luciferase in transgenic mice: Regulatory studies in vivo and in vitro}, volume={142}, ISSN={["0013-7227"]}, DOI={10.1210/en.142.6.2260}, abstractNote={Transgenic mice harboring the ovine FSHβ (oFSHβ) promoter plus first intron (from −4741 to +759 bp) linked to a luciferase reporter gene (oFSHβLuc) were generated to determine whether this promoter can direct tissue-specific expression in vivo and serve as a model for studying hormonal regulation of the FSHβ gene. Of six lines of transgenic mice analyzed, luciferase was detected uniquely in the pituitaries of five of them. Pituitary luciferase activity was decreased 51–99% by chronic GnRH treatment (Lupron depot). Orchidectomy caused a 2- to 8-fold increase, and ovariectomy caused a 2- to 27-fold increase in pituitary luciferase activity. Furthermore, pituitary luciferase expression was consistently higher on estrus than on diestrus (3- to 20-fold). These data strongly suggested that the transgene was expressed specifically in pituitary gonadotropes and regulated in the same way as the endogenous mouse FSHβ gene. Using primary pituitary cell cultures prepared from these transgenic mice, basal luciferase expression was maximal on day 3 and then decreased by day 6 of culture, a pattern reflected by endogenous mouse FSH secretion. In these pituitary cultures, basal oFSHβLuc expression was decreased 61–82% by follistatin or 59–79% by inhibin. Similarly, mouse FSH secretion was decreased 71% by follistatin or 65% by inhibin. Progesterone inhibited oFSHβLuc expression by 44–51%, but it had no effect on endogenous mouse FSH secretion. Estradiol lowered FSH secretion by 21%, but did not decrease oFSHβLuc expression significantly. In conclusion, these data demonstrated the ability of the oFSHβ promoter to direct expression of a reporter gene specifically to pituitary gonadotropes in transgenic mice. Studying oFSHβLuc expression in vivo and in cell cultures derived from pituitaries of these transgenic mice should prove useful for understanding many features of FSHβ regulation.}, number={6}, journal={ENDOCRINOLOGY}, author={Huang, HJ and Sebastian, J and Strahl, BD and Wu, JC and Miller, WL}, year={2001}, month={Jun}, pages={2260–2266} }
 @article{huang_sebastian_strahl_wu_miller_2001, title={Transcriptional regulation of the ovine follicle-stimulating hormone-beta gene by activin and gonadotropin-releasing hormone (GnRH): Involvement of two proximal activator protein-1 sites for GnRH stimulation}, volume={142}, ISSN={["0013-7227"]}, DOI={10.1210/en.142.6.2267}, abstractNote={Previous studies from our laboratory demonstrated that a transgene consisting of the promoter for the ovine FSH β-subunit gene and a luciferase reporter (wt-oFSHβLuc) was expressed and regulated like the FSHβ gene in vivo and in vitro. In the present study pituitary cultures were prepared from these transgenic mice as well as mice carrying mutated oFSHβLuc lacking two functional activator protein-1 (AP-1) sites at −120 and −83 bp (mut-oFSHβLuc). These AP-1 sites were reported necessary for induction of oFSHβLuc by GnRH in a HeLa cell system. To examine the importance of the two AP-1 sites in mediating GnRH and activin effects in primary gonadotropes, pituitary cultures derived from transgenic mice were pretreated with follistatin to remove activin or activin-like factors present in the cultures. Follistatin lowered luciferase expression in cultures carrying both wt-oFSHβLuc and mut-oFSHβLuc transgenes by 74–86%, and subsequent addition of activin induced luciferase expression of both wt- and mut-transgenes by 4- to 14-fold within 4 h, suggesting that these AP-1 sites are not involved in activin stimulation of FSHβ gene transcription. When GnRH was added along with activin, the wt-oFSHβLuc transgene was induced 200% compared with activin alone, but this effect was not observed with the mut-oFSHβLuc transgene. These data confirmed the HeLa cell studies showing that GnRH signals through two AP-1 sites to increase oFSHβ transcription in gonadotropes. However, as the mutation of both AP-1 sites had no apparent effect on the expression and regulation of the transgene in vivo (basal, castration, GnRH down-regulation, cycle stage, and GnRH immunoneutralization), it appears that these AP-1 sites have little influence over the major effect of GnRH observed in vivo. These data also showed that activin plays a major role in transcriptional regulation of the FSHβ gene, and the oFSHβ promoter contains the activin response element(s) that is as yet undefined.}, number={6}, journal={ENDOCRINOLOGY}, author={Huang, HJ and Sebastian, J and Strahl, BD and Wu, JC and Miller, WL}, year={2001}, month={Jun}, pages={2267–2274} }
 @article{gardner_sebastian_miller_2000, title={Estradiol induces and hyperglycosylates the receptor for ovine gonadotropin-releasing hormone}, volume={141}, ISSN={["1945-7170"]}, DOI={10.1210/en.141.1.91}, abstractNote={The crucial first link between GnRH and its pleiotropic stimulation of the reproductive system is its receptor (GnRHRec). In mammals, 17beta-estradiol is a major regulator of GnRH action, and part of its regulation occurs at the level of the GnRHRec. In ovine pituitary cultures, estradiol simultaneously increases GnRHRec and GnRH-stimulated LH secretion (the LH response), but after 6-15 h the effect of estradiol becomes paradoxical, and the LH response rapidly decreases to control levels (by 24 h), whereas GnRHRec remains elevated. A preliminary study used photoaffinity labeling of the GnRHRec to show that estradiol can induce 38- and 43-kDa GnRHRec. The photoaffinity technique has been used here to 1) further investigate estradiol-mediated induction of GnRHRec, 2) define the nature of the different sized GnRHRecs, and 3) determine whether the larger size is related to degradation of the LH response. The effect of estradiol is compared with that of inhibin, which only induces the 38-kDa GnRHRec and always increases the LH response to GnRH treatment. Receptors for GnRH in ovine pituitary cultures were photoaffinity labeled with [125I](azidobenzoyl-D-Lys6-des-Gly10)-GnRH-N-ethylamide and analyzed by SDS-PAGE. Treatment with estradiol or inhibin for 6-24 h induced a 38-kDa GnRHRec only. Further treatment with estradiol (>24 h), but not inhibin, shifted the apparent Mr of the GnRHRec to 43 kDa. Phosphatase treatment did not reverse this apparent Mr change. Analysis of receptor glycosylation using N-glycosidase F or tunicamycin showed that the 43-kDa GnRHRec was a hyperglycosylated form of the 38-kDa GnRHRec. The 38-kDa GnRHRec, in turn, was a glycosylated form of the 29-kDa GnRHRec. The studies presented here define several glycosylated intermediates of the ovine GnRHRec that are induced by estradiol and/or inhibin. The function of estrogen-mediated hyperglycosylation is unclear, but kinetic studies dissociate it from degeneration of the LH response to GnRH.}, number={1}, journal={ENDOCRINOLOGY}, author={Gardner, DB and Sebastian, J and Miller, WL}, year={2000}, month={Jan}, pages={91–99} }
 @article{robinette_wada_arroll_levy_miller_noga_1998, title={Antimicrobial activity in the skin of the channel catfish Ictalurus punctatus: characterization of broad-spectrum histone-like antimicrobial proteins}, volume={54}, ISSN={["1420-9071"]}, DOI={10.1007/s000180050175}, abstractNote={Three antibacterial proteins were isolated from acid extracts of channel catfish (Ictalurus punctatus) skin by cation exchange chromatography and reverse-phase high-pressure liquid chromatography. The molecular masses of the proteins were 15.5, 15.5 and 30 kD as determined by SDS-polyacrylamide gel electrophoresis. Mass spectrometry, amino acid composition and amino acid sequence data suggest that the most abundant protein is closely related to histone H2B. The H2B-like protein was inhibitory to Aeromonas hydrophila and Saprolegnia spp., which are important bacterial and fungal pathogens of fish. These findings suggest that histones may be important defensive molecules in fish.}, number={5}, journal={CELLULAR AND MOLECULAR LIFE SCIENCES}, author={Robinette, D and Wada, S and Arroll, T and Levy, MG and Miller, WL and Noga, EJ}, year={1998}, month={May}, pages={467–475} }
 @article{strahl_huang_sebastian_ghosh_miller_1998, title={Transcriptional activation of the ovine follicle-stimulating hormone beta-subunit gene by gonadotropin-releasing hormone: Involvement of two activating protein-1-binding sites and protein kinase C}, volume={139}, ISSN={["1945-7170"]}, DOI={10.1210/en.139.11.4455}, number={11}, journal={ENDOCRINOLOGY}, author={Strahl, BD and Huang, HJ and Sebastian, J and Ghosh, BR and Miller, WL}, year={1998}, month={Nov}, pages={4455–4465} }
 @article{childs_miller_miller_1997, title={Differential effects of inhibin on gonadotropin stores and gonadotropin-releasing hormone binding to pituitary cells from cycling female rats}, volume={138}, ISSN={["0013-7227"]}, DOI={10.1210/en.138.4.1577}, abstractNote={Numerous studies of rat pituitaries have reported that inhibin suppresses the synthesis and release of FSH and decreases the release of LH. The latter effect seems to be related to the down-regulation of receptors for GnRH. The studies reported here identified cellular changes behind the inhibitory effects of inhibin on gonadotropes to learn whether its effects are mediated by changes in subtypes of gonadotropes. Cell populations from diestrous day 2 and proestrous (morning) rats were collected, dispersed to single cell populations, and plated in medium containing either recombinant 32-kDa inhibin or porcine follicular fluid for 24 h. GnRH binding was detected by exposing the cells to a biotinylated analog (Bio-GnRH) for 10 min before fixation, followed by avidin-peroxidase labeling protocols to detect the biotin on the analog. In parallel fields, the cells were further identified by immunolabeling for LH or FSH β-subunits or for GH with a different colored reaction product. The most striking changes were seen in cells from proestrous rats. Inhibin reduced the percentages of Bio-GnRH target cells in the population by 60% and the area and density of Bio-GnRH label on the remaining cells. Inhibin reduced the percentages of FSH cells by 30% and caused nearly a 60% reduction in the binding of Bio-GnRH by this cell type (from 83% of FSH cells to 32% of FSH cells). Inhibin also reduced the area of FSH cells and the density of FSH stores. Inhibin’s effects on LH cells were limited to a reduction in the area of the cells and the density of LH stores, but not the number of LH cells. In addition, it reduced the percentages of LH cells with Bio-GnRH receptors from 84% to 40%. When cells with GH were analyzed, inhibin had no effect on their percentages, areas, or GH stores. In populations from proestrous rats, inhibin reduced the percentages of GH cells with Bio-GnRH binding from 38% to 21%. These data suggest that inhibin’s target cell is the abundant multihormonal gonadotrope that contains LH, FSH, and GH and predominates during proestrus. Inhibin’s effects are most severe on FSH cells, which suggests that it may either selectively affect FSH synthesis and stores in bihormonal gonadotropes and/or affect monohormonal FSH cells. Thus, mechanisms behind its inhibitory effects include 1) a reduction in the percentage of Bio-GnRH target cells, 2) a reduction in the area of Bio-GnRH-binding sites on individual cells, and 3) a reduction in the stores of FSH and the percentages of FSH cells. These last effects are consistent with known reductions in FSH synthesis. The effects of inhibin on LH secretion may be secondary to the effects on Bio-GnRH receptors in bihormonal gonadotropes.}, number={4}, journal={ENDOCRINOLOGY}, author={Childs, GV and Miller, BT and Miller, WL}, year={1997}, month={Apr}, pages={1577–1584} }
 @article{strahl_huang_pedersen_wu_ghosh_miller_1997, title={Two proximal activating protein-1-binding sites are sufficient to stimulate transcription of the ovine follicle-stimulating hormone-beta gene}, volume={138}, ISSN={["1945-7170"]}, DOI={10.1210/en.138.6.2621}, abstractNote={FSH is an important regulator of mammalian gametogenesis and the female reproductive cycle. Although little is known about the transcriptional regulation of the β-subunit (the rate-limiting subunit of FSH synthesis), sequence analysis of the ovine FSHβ promoter has revealed a number of potential activating protein-1 (AP-1; Jun/Fos)-binding sites. To determine whether the gene encoding theβ -subunit of ovine FSH (oFSHβ) is responsive to AP-1 transcriptional complexes, chimeric constructs containing deleted portions of the oFSHβ promoter fused to a luciferase reporter were transiently transfected along with c-Jun and c-Fos expression constructs into JAR cells. Analysis of these deletion constructs revealed that the proximal promoter of oFSHβ is highly stimulated by c-Jun and c-Fos proteins (typically 20-fold with a reporter construct containing oFSHβ sequences from −215 to +759 bp). This stimulation was lost when a similar construct containing sequences from −84 to+ 759 bp was tested. Transcriptional start site analysis using reverse transcription-PCR verified that the transcriptional initiation of the− 215-bp deletion construct, with or without cotransfected c-Jun/c-Fos, was the same as that observed in vivo. Computer analysis of oFSHβ sequences from −215 to +1 bp identified four putative AP-1-like elements, located at −155, −120, −83, and −10 bp. Gel retardation experiments using oligonucleotides corresponding to the four putative AP-1-like sites revealed that only −120 and −83 sites in oFSHβ bound AP-1 proteins in vitro. Site-directed mutagenesis of the −120 and −83 sites showed that each element was required for stimulation by c-Jun and c-Fos proteins as well as 12-O-tetradecanoyl phorbol-13-acetate in transient transfection assays. Finally, immunocytochemical dual labeling was used to show that more than 75% of all FSHβ-containing cells in ovine pituitary sections from cycling ewes contained nuclear c-Jun, JunB, JunD, and Fos proteins. These data, taken together, show that oFSHβ transcription can be stimulated by c-Jun and c-Fos proteins via two functionally linked AP-1-like sites in the oFSHβ proximal promoter and that these sites are likely to be important regulators of FSH production in vivo.}, number={6}, journal={ENDOCRINOLOGY}, author={Strahl, BD and Huang, HJ and Pedersen, NR and Wu, JC and Ghosh, BR and Miller, WL}, year={1997}, month={Jun}, pages={2621–2631} }
 @inbook{miller_1993, title={Regulation of pituitary gonadotropins by gonadotropin-releasing hormone, estradiol, progesterone, inhibin, and activin}, ISBN={0471561460}, booktitle={Genes in mammalian reproduction}, publisher={New York : Wiley-Liss}, author={Miller, W. L.}, year={1993}, pages={247} }
 @article{miller_huang_1985, title={SECRETION OF OVINE LUTEINIZING-HORMONE INVITRO - DIFFERENTIAL POSITIVE CONTROL BY 17-BETA-ESTRADIOL AND A PREPARATION OF PORCINE OVARIAN INHIBIN}, volume={117}, ISSN={["1945-7170"]}, DOI={10.1210/endo-117-3-907}, abstractNote={17 beta-Estradiol (E2) and an enriched preparation of porcine ovarian inhibin can positively regulate LH secretion in two different ways in ovine pituitary cell cultures. One major effect of these agents is to increase the sensitivity of cultures to low levels of LHRH or LHRH analogs. The second effect is to increase culture responsiveness to high levels of LHRH or its analogs. Prolonged treatment with 1 nM E2, for instance, increased pituitary sensitivity to D-Lys6-LHRH by 10-fold, since it lowered the ED50 of D-Lys6-LHRH from approximately 400 pM to about 40 pM; the maximum LH secretory response to D-Lys6-LHRH (1-100 nM) was not changed, however. In contrast, short term treatment with an E2-free preparation of porcine ovarian inhibin could increase, by 350%, normal LH responsiveness to high levels of D-Lys6-LHRH (1-100 nM), but had no effect on sensitivity; the ED50 remained unchanged. Chronic treatment with inhibin eventually increased sensitivity, and interestingly, a short treatment with E2 (6 h) caused a major increase in both sensitivity and responsiveness, but the responsiveness component usually disappeared between 6 and 48 h of chronic E2 treatment. Treatment with both E2 and the inhibin preparation caused the greatest increase in pituitary sensitivity to D-Lys6-LHRH (ED50 decreased to approximately 30 pM), but the increased responsiveness to D-Lys6-LHRH (1-100 nM) declined with time as with E2-only treatment. The different actions of E2 and the ovarian inhibin preparation seen to perturb different aspects of LH secretion. Therefore, study of these phenomena may uncover novel molecular details governing LHRH-stimulated LH release. Furthermore, these two types of LH secretory control may be physiologically important, at least in sheep, and perhaps in other animals as well.}, number={3}, journal={ENDOCRINOLOGY}, author={MILLER, WL and HUANG, ESR}, year={1985}, pages={907–911} }
 @article{miller_huang_1981, title={ANTI-ESTROGENS AND OVINE GONADOTROPHS - ANTAGONISM OF ESTROGEN-INDUCED CHANGES IN GONADOTROPIN SECRETIONS}, volume={108}, ISSN={["0013-7227"]}, DOI={10.1210/endo-108-1-96}, abstractNote={17 beta-Estradiol (E2) was found to affect gonadotrophs in long term ovine pituitary cell culture in several ways which were easily measured. It inhibited basal FSH secretion and, concomitantly, stimulated LH secretion. High physiological levels of E2 (10(-10) M; 27 pg/ml) were able to elicit about 70% of the maximal responses, and the effects were developed fully between 6-24 h after the addition of E2. In culture, the antiestrogens (AEs) tamoxifen, nafoxidine, and CI-680 antagonized both the inhibitory and stimulatory effects of E2 (10(-10) M) on gonadotropin secretion in a dose-dependent manner, and 10(-6) M AE was fully effective. At concentrations of 10(-6) M or lower, the AEs showed no significant intrinsic estrogenicity. At higher concentrations, the AEs progressively altered gonadotropin secretion in an apparently nonspecific fashion. Preincubations of cultures with AEs (3.3 X 10(-6) M) antagonized E2 action (10(-10) M) for approximately 18-24 h after the removal of AEs. Tamoxifen appeared to be the most potent AE tested. In conclusion, long term ovine pituitary cell cultures have provided a useful in vitro system for testing the biological potencies of AE analogs. AEs appeared to act as pure estrogen antagonists in ovine gonadotrophs.}, number={1}, journal={ENDOCRINOLOGY}, author={MILLER, WL and HUANG, ESR}, year={1981}, pages={96–102} }