Search

Mammalian ovarian follicular development and maturation of an oocyte competent to be fertilized and develop into an embryo depends on tightly regulated, spatiotemporally orchestrated crosstalk among cell death, survival, and differentiation signals through extra- and intraovarian signals, as well as on a permissive ovarian follicular microenvironment. Neuregulin-1 (NRG1) is a member of the epidermal growth factor–like factor family that mediates its effects by binding to a member of the erythroblastoma (ErbB) family. Our experimental results suggest gonadotropins promote differential expression of NRG1 and erbB receptors in granulosa cells (GCs), and NRG1 in theca cells during follicular development, and promote NRG1 secretions in the follicular fluid (FF) of rat ovaries. During the estrous cycle of rat, NRG1 and erbB receptors are differentially expressed in GCs and correlate positively with serum gonadotropins and steroid hormones. Moreover, in vitro experimental studies suggest that the protein kinase C inhibitor staurosporine (STS) causes the physical destruction of GCs by the activation of caspase-3. Exogenous NRG1 treatment of GCs delayed onset of STS-induced apoptosis and inhibited cleaved caspase-3 expressions. Moreover, exogenous NRG1 treatment of GCs alters STS-induced death by maintaining the expression of ErbB2, ErbB3, pAkt, Bcl2, and BclxL proteins. Taken together, these studies demonstrate that NRG1 is gonadotropin dependent, differentially regulated in GCs and theca cells, and secreted in ovarian FF as an intracellular survival factor that may govern follicular maturation.

Background We determined influenza A(H1N1)pdm09 antibody levels before and after the first wave of the pandemic in an urban community in Dhaka, Bangladesh. Methods We identified a cohort of households by stratified random sampling. We collected baseline serum specimens during July-August 2009, just prior to the initial wave of the 2009 pandemic in this community and a second specimen during November 2009, after the pandemic peak. Paired sera were tested for antibodies against A(H1N1)pdm09 virus using microneutralization assay and hemagglutinin inhibition (HI) assay. A fourfold increase in antibody titer by either assay with a titer of ≥40 in the convalescent sera was considered a seroconversion. At baseline, an HI titer of ≥40 was considered seropositive. We collected information on clinical illness from weekly home visits. Results We tested 779 paired sera from the participants. At baseline, before the pandemic wave, 1% overall and 3% of persons >60 years old were seropositive. After the first wave of the pandemic, 211 (27%) individuals seroconverted against A(H1N1)pdm09. Children aged 5-17 years had the highest proportion (37%) of seroconversion. Among 264 (34%) persons with information on clinical illness, 191 (72%) had illness >3 weeks prior to collection of the follow-up sera and 73 (38%) seroconverted. Sixteen (22%) of these 73 seroconverted participants reported no clinical illness. Conclusion After the first pandemic wave in Dhaka, one in four persons were infected by A(H1N1)pdm09 virus and the highest burden of infection was among the school-aged children. Seroprevalence studies supplement traditional surveillance systems to estimate infection burden.

Seroprevalence of antibodies to influenza A(H1N1)pdm09 virus among 193 emergency department health care personnel was similar among 147 non-health care personnel (odds ratio 1.4, 95% CI 0.8-2.4). Working in an acute care setting did not substantially increase risk for virus infection above risk conferred by community-based exposures.

Background During April–July 2009, U.S. hospitalization rates for 2009 pandemic influenza A (H1N1) virus (H1N1pdm09) infection were estimated at 4·5/100 000 persons. We describe rates and risk factors for H1N1pdm09 infection among American Indians (AIs) in four isolated southwestern U.S. communities served by the Indian Health Service (IHS). Methods We reviewed clinical and demographic information from medical records of AIs hospitalized during May 1–July 21, 2009 with severe acute respiratory infection (SARI). Hospitalization rates were determined using denominator data provided by IHS. H1N1pdm09 infection was confirmed with polymerase chain reaction, rapid tests, or convalescent serology. Risk factors for more severe (SARI) versus milder [influenza-like illness (ILI)] illness were determined by comparing confirmed SARI patients with outpatients with ILI. Results Among 168 SARI-hospitalized patients, 52% had confirmed H1N1pdm09 infection and 93% had >1 high-risk condition for influenza complications. The H1N1pdm09 SARI hospitalization rate was 131/100 000 persons [95% confidence interval (CI), 102–160] and was highest among ages 0–4 years (353/100 000; 95% CI, 215–492). Among children, asthma (adjusted odds ratio [aOR] 3·2; 95% CI, 1·2–8·4) and age

Avoiding or limiting contact with swine at agricultural events may help prevent A(H3N2)v virus infections in such settings., During August 2011, influenza A (H3N2) variant [A(H3N2)v] virus infection developed in a child who attended an agricultural fair in Pennsylvania, USA; the virus resulted from reassortment of a swine influenza virus with influenza A(H1N1)pdm09. We interviewed fair attendees and conducted a retrospective cohort study among members of an agricultural club who attended the fair. Probable and confirmed cases of A(H3N2)v virus infection were defined by serology and genomic sequencing results, respectively. We identified 82 suspected, 4 probable, and 3 confirmed case-patients who attended the fair. Among 127 cohort study members, the risk for suspected case status increased as swine exposure increased from none (4%; referent) to visiting swine exhibits (8%; relative risk 2.1; 95% CI 0.2–53.4) to touching swine (16%; relative risk 4.4; 95% CI 0.8–116.3). Fairs may be venues for zoonotic transmission of viruses with epidemic potential; thus, health officials should investigate respiratory illness outbreaks associated with agricultural events.

Ceramide is a key factor in inducing germ cell apoptosis by translocating from cumulus cells into the adjacent oocyte and lipid rafts through gap junctions. Therefore studies designed to elucidate the mechanistic pathways in ceramide induced granulosa cell (GC) apoptosis and follicular atresia may potentially lead to the development of novel lipid-based therapeutic strategies that will prevent infertility and premature menopause associated with chemo and/or radiation therapy in female cancer patients. Our previous studies have shown that Prohibitin (PHB) is intimately involved in GCs differentiation, atresia, and luteolysis.In the present study, we have examined the functional effects of loss-/gain-of-function of PHB using adenoviral technology in delaying apoptosis induced by the physiological ligand ceramide in rat GCs.Under these experimental conditions, exogenous ceramide C-8 (50 μM) augmented the expression of mitochondrial PHB and subsequently cause the physical destruction of GC by the release of mitochondrial cytochrome c and activation of caspase-3. In further studies, silencing of PHB expression by adenoviral small interfering RNA (shRNA) sensitized GCs to ceramide C8-induce apoptosis. In contrast, adenovirus (Ad) directed overexpression of PHB in GCs resulted in increased PHB content in mitochondria and delayed the onset of ceramide induced apoptosis in the infected GCs.Taken together, these results provide novel evidences that a critical level of PHB expression within the mitochondria plays a key intra-molecular role in GC fate by mediating the inhibition of apoptosis and may therefore, contribute significantly to ceramide induced follicular atresia.

Prohibitin is a ubiquitous and highly conserved protein implicated as an important regulator in cell survival. Prohibitin content is inversely associated with cell proliferation, but it increases during granulosa cell differentiation as well as in earlier events of apoptosis in a temperature-sensitive granulosa cell line. In the present study, we have characterized the spatial expression patterns for prohibitin using established in vivo models for the induction of follicular development and atresia in the mammalian ovary. Comparative Western blot analyses of granulosa cell lysates from control ovaries and from ovaries primed with eCG or treated with eCG plus anti-eCG (gonadotropin withdrawal) were conducted. Prohibitin was immunolocalized in rat ovarian sections probed with antibodies against either proliferating cell nuclear antigen (PCNA) or cholesterol side-chain cleavage cytochrome P450 (P450(scc)) or in terminal deoxynucleotidyl transferase-mediated dUTP nick end labeled sections. Additionally, porcine oocytes, zygotes, and blastocyts were also immunolocalized with prohibitin antibody. Immunolocalization revealed the presence of prohibitin in granulosa cells, theca-interstitial cells, and the oocyte. The results indicate that prohibitin protein expression in the gonadotropin-treated cells was upregulated. Immunoreactivity of prohibitin was inversely related to PCNA expression during follicular maturation and colocalized with P450(scc). Prohibitin appeared to be translocated from the cytoplasm to the nucleus in atretic follicles, germinal vesicle-stage oocytes, zygotes, and blastocysts. These results suggest that prohibitin has several functional regulatory roles in granulosa and theca-interstitial cells and in the ovum during follicular maturation and atresia. It is likely that prohibitin may play an important role in determining the fate of these cells and eventual follicular destiny.

Prohibitin is an evolutionary conserved protein that is associated with cellular differentiation, atresia, and luteolysis in the rat ovary. However, the specific cellular location and function of prohibitin in ovarian cells has not been clearly elucidated. To characterize the expression of prohibitin during cell proliferation, differentiation, and cell death, we have successfully established a temperature-sensitive granulosa cell line, designated RGA-1. At a permissive temperature of 33 C, RGA-1 cells proliferate, but revert to a differentiated phenotype at a nonpermissive temperature of 39 C. Significant inductions of prohibitin mRNA and protein expression were observed in the differentiated phenotype when compared with proliferating cells. Differentiated RGA-1 cells were found to express inhibin alpha- and beta-transcripts, as well as steroidogenic acute regulatory protein and peripheral-type benzodiazepine receptor proteins in a manner reminiscent of steroidogenic functional responses observed in primary differentiated granulosa cells. Prohibitin expression correlated well with the expression of these steroidogenic proteins. At 39 C, RGA-1 cells also displayed increases in p53 protein levels, indicative of growth arrest in the nonproliferating cells. Confocal and electron microscopic examinations revealed increased prohibitin localization to the mitochondria at 39 C, along with changes in mitochondrial size and shape. These changes were accompanied by marked reductions in cytochrome c oxidase subunit II levels and in unit mitochondrial transmembrane potential. In addition, cell fractionation studies demonstrated that the prohibitin protein was mainly localized to the mitochondrial membrane. Collectively, these findings suggest a role for prohibitin in mitochondrial structure and function during growth and differentiation in ovarian granulosa cells. Prohibitin expression may also be indicative of mitochondrial destabilization during apoptosis-related events.