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Hepatitis C Virus (HCV) is a major public health concern, with no effective vaccines currently available and 3% of the world's population being infected. Despite the existence of both B- and T-cell immunity in HCV-infected patients, chronic viral infection and HCV-related malignancies progress. Here we report the identification of a novel HCV TCR from an HLA-A2-restricted, HCV NS3:1073-1081-reactive CTL clone isolated from a patient with chronic HCV infection. We characterized this HCV TCR by expressing it in human T cells and analyzed the function of the resulting HCV TCR-transduced cells. Our results indicate that both the HCV TCR-transduced CD4(+) and CD8(+) T cells recognized the HCV NS3:1073-1081 peptide-loaded targets and HCV(+) hepatocellular carcinoma cells (HCC) in a polyfunctional manner with cytokine (IFN-gamma, IL-2, and TNF-alpha) production as well as cytotoxicity. Tumor cell recognition by HCV TCR transduced CD8(-) Jurkat cells and CD4(+) PBL-derived T cells indicated this TCR was CD8-independent, a property consistent with other high affinity TCRs. HCV TCR-transduced T cells may be promising for the treatment of patients with chronic HCV infections.

A commitment to diversity, equity, inclusion, and belonging in medical education requires addressing both explicit and implicit biases based on sexual orientation, gender identity and expression, and sex characteristics and the intersectionality with other identities. Heterosexism and heteronormative attitudes contribute to health and healthcare disparities for lesbian, gay, bisexual, transgender and queer or questioning, intersex, asexual individuals. Student, trainee, and faculty competencies in medical education curricula regarding the care of lesbian, gay, bisexual, transgender and queer or questioning, intersex, asexual patients and those who are gender nonconforming or born with differences of sex development allow for better understanding and belonging within the clinical learning environment of lesbian, gay, bisexual, transgender and queer/questioning, intersex, asexual learners and educators. The Association of Professors of Gynecology and Obstetrics issued a call to action to achieve a future free from racism and bias through inclusivity in obstetrics and gynecology education and healthcare, which led to the creation of the Association of Professors of Gynecology and Obstetrics Diversity, Equity, and Inclusion Guidelines Task Force. The task force initially addressed racism, racial- and ethnicity-based bias, and discrimination in medical education and additionally identified other groups that are subject to bias and discrimination, including sexual orientation, gender identity and expression, and sex characteristic identities, persons with disabilities, and individuals with various religious and spiritual practices. In this scholarly perspective, the authors expand on previously developed guidelines to address sexual orientation, gender identity and expression, and sex characteristics bias, heterosexism, and heteronormative attitudes in obstetrics and gynecology educational products, materials, and clinical learning environments to improve access and equitable care to vulnerable individuals of the lesbian, gay, bisexual, transgender and queer or questioning, intersex, asexual community., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Objective  This study aimed to identify factors associated with meeting the Obstetric Care Consensus (OCC) guidelines for nulliparous, term, singleton, and vertex (NTSV) cesarean births. Materials and methods  This was a retrospective case control study of women with NTSV cesarean births between January 2014 and December 2017 at single tertiary care center. Demographics and clinical characteristics were compared between women with NTSV cesarean births which did or did not meet OCC guidelines. A multivariable logistic regression model was used to evaluate the effect of each variable on the odds of meeting OCC guidelines. Results  There were 1,834 women with NTSV cesarean births of which 744 (40.6%) met OCC guidelines for delivery and 1,090 (59.4%) did not. After controlling for confounding factors, the odds of meeting OCC guidelines were increased for in-house providers managing with residents (adjusted odds ratio [aOR] = 2.03, 95% confidence interval [CI]: 1.44-2.87) and without residents (aOR = 1.66, 95% CI: 1.30-2.12), compared with non-in-house providers managing without residents. There was no significant difference in the odds of meeting OCC guidelines for in-house providers managing with or without residents (aOR = 1.23, 95% CI: 0.84-1.79). Conclusion  After adjusting for confounding factors, in-house provider coverage, regardless of resident involvement, is associated with increased odds of NTSV cesarean births meeting OCC guidelines. Key Points Frequency of adherence to OCC guidelines for NTSV cesarean births was 40.6%.Neither patient demographics nor comorbidities was associated with the odds of meeting OCC guidelines.In-house providers are associated with increased odds of meeting OCC guidelines., Competing Interests: Conflict of Interest The authors report no financial or commercial conflicts of interest. This study did not receive any funding from external or internal sources., (The Author(s). This is an open access article published by Thieme under the terms of the Creative Commons Attribution-NonDerivative-NonCommercial License, permitting copying and reproduction so long as the original work is given appropriate credit. Contents may not be used for commercial purposes, or adapted, remixed, transformed or built upon. ( https://creativecommons.org/licenses/by-nc-nd/4.0/ ).)

Purpose: We conducted a randomized phase III trial to determine whether adjuvant chemotherapy improves survival in women with uterine leiomyosarcoma., Methods: Women with uterus-confined, high-grade leiomyosarcoma who were confirmed disease free by imaging were randomly assigned to four cycles of gemcitabine plus docetaxel, followed by four cycles of doxorubicin, or to observation. All were followed for evidence of recurrence. The primary end point was overall survival (OS)., Results: With international collaboration, 38 of the targeted accrual of 216 patients were enrolled, after which the study was closed by the National Cancer Institute for accrual futility. Twenty patients were assigned to chemotherapy, 18 to observation. Among the 17 patients treated with at least one cycle of chemotherapy, grade 3 or 4 toxicities were observed in 47%; among the 18 patients assigned to observation, one had grade 3 hypertension. There were six deaths (chemotherapy, n = 5; observation, n = 1), all due to disease. The restricted mean survival time for OS was estimated as 34.3 months (95% CI, 25.3 to 43.3 months) in the chemotherapy arm and as 46.4 months (95% CI, 43.6 to 49.1 months) in the observation arm. There were eight recurrences in each arm. The restricted mean survival time for recurrence-free survival was estimated as 18.1 (95% CI, 14.2 to 22.0) months in the chemotherapy arm and as 14.6 months (95% CI, 10.3 to 19.0 months) in the observation arm. Neither survival outcome comparison was considered statistically robust, due to the small sample size., Conclusion: Despite international collaboration to test the role of adjuvant chemotherapy in uterine-confined leiomyosarcoma, this study was closed for accrual futility. Although the sample size precludes robust statistical comparison, observed OS and recurrence-free survival data do not show superior outcomes with adjuvant chemotherapy.

Marijuana use was recently reported to have a positive cross-sectional association with human papillomavirus (HPV)-related head and neck cancer. Laboratory data suggest that marijuana could have an immunomodulatory effect. Little is known, however, regarding the effects of marijuana use on cervical HPV or neoplasia. Therefore, we studied the natural history (i.e., prevalence, incident detection, clearance/persistence) of cervical HPV and cervical neoplasia (i.e., squamous intraepithelial lesions; SIL) in a large prospective cohort of 2,584 HIV-seropositive and 915 HIV-seronegative women. Marijuana use was classified as ever/never, current/not current, and by frequency and duration of use. No positive associations were observed between use of marijuana, and either cervical HPV infection or SIL. The findings were similar among HIV-seropositive and HIV-seronegative women, and in tobacco smokers and nonsmokers. These data suggest that marijuana use does not increase the burden of cervical HPV infection or SIL.

High serum levels of insulin-like growth factor-I (IGF-I) are reported to be a risk factor for several common cancers, and recent cross-sectional data suggest a possible additional association of IGF-I with cervical neoplasia. To prospectively assess whether circulating IGF-I levels influence the natural history of oncogenic human papillomavirus (HPV), the viral cause of cervical cancer, we conducted a pilot investigation of 137 women who underwent semiannual type-specific HPV DNA PCR testing and cervical cytology. Total IGF-I and IGF binding protein-3 (IGFBP-3), the most abundant IGFBP in circulation, were measured using baseline serum specimens. Having a high IGF-I/IGFBP-3 ratio was associated with increased persistence of oncogenic HPV infection [that is, a lower rate of clearance; adjusted hazard ratio (AHR), 0.14; 95% confidence interval (95% CI), 0.04-0.57], whereas IGFBP-3 was inversely associated with both the incident detection of oncogenic HPV (AHR, 0.35; 95% CI, 0.13-0.93) and the incidence of oncogenic HPV-positive cervical neoplasia (that is, squamous intraepithelial lesions at risk of progression; AHR, 0.07; 95% CI, 0.01-0.66). These prospective data provide initial evidence that the IGF axis may influence the natural history of oncogenic HPV.

Objective: To compare Müllerian inhibiting substance (MIS) levels in serum obtained during the early follicular phase to those obtained randomly during the menstrual cycle. To determine whether HIV infection influences early follicular MIS levels, an early marker of ovarian aging., Design: A cross-sectional study., Setting: Women's Interagency HIV Study, a multicenter prospective study., Patient(s): Serum samples obtained from 263 (187 HIV infected and 76 uninfected) participants of the Women's Interagency HIV Study who reported menstrual bleeding during the preceding 6 months and who were not taking exogenous hormones., Intervention(s): Early follicular (cycle days 2-5) MIS samples were compared with serum samples that had been obtained without regard to menstrual cycle phase. Comparison samples were obtained within 6 weeks before or within 3 to 6 months after the early follicular samples. Early follicular FSH, E(2), inhibin B, and MIS levels were also compared between the HIV infected and uninfected women., Main Outcome Measure(s): Correlation between early follicular MIS and prior and subsequent samples. Comparison of serum markers of ovarian reserve between HIV positive and negative women., Result(s): The MIS values from early follicular and other random cycle phases were highly correlated with each other (r > 0.93). In multivariate analysis, increased age and FSH level and lower inhibin B levels were associated with lower MIS level; MIS values did not vary by HIV serostatus., Conclusion(s): Without regard to cycle phase, MIS was similar during early follicular phase and highly correlated with early follicular FSH and inhibin B in women with and without HIV. Measurement of serum MIS offers a simplified method of determining ovarian reserve using specimens obtained without menstrual phase timing. Furthermore, using biologic measures of reproductive aging, we found no evidence that HIV infection influences ovarian aging.

Objective: To characterize ovarian failure and prolonged amenorrhea from other causes in women who are both human immunodeficiency virus (HIV) seropositive and seronegative., Methods: This was a cohort study nested in the Women's Interagency HIV Study, a multicenter U.S. study of HIV infection in women. Prolonged amenorrhea was defined as no vaginal bleeding for at least 1 year. A serum follicle stimulating hormone more than 25 milli-International Units/mL and prolonged amenorrhea were used to define ovarian failure. Logistic regressions, chi2, and t tests were performed to estimate relationships between HIV-infection and cofactors with both ovarian failure and amenorrhea from other causes., Results: Results were available for 1,431 women (1,139 HIV seropositive and 292 seronegative). More than one half of the HIV positive women with prolonged amenorrhea of at least 1 year did not have ovarian failure. When adjusted for age, HIV seropositive women were about three times more likely than seronegative women to have prolonged amenorrhea without ovarian failure. Body mass index, serum albumin, and parity were all negatively associated with ovarian failure in HIV seropositive women., Conclusion: HIV serostatus is associated with prolonged amenorrhea. It is difficult to ascertain whether the cause of prolonged amenorrhea is ovarian in HIV-infected women without additional testing., Level of Evidence: II-2.

Objectives: To evaluate changes over time in rates of bacterial vaginosis (BV), trichomoniasis (TV), and yeast vaginitis (YV) among HIV-infected and similar HIV-uninfected women., Methods: Two thousand fifty-six HIV-infected women and 554 HIV-uninfected women were evaluated semiannually from 1994 until March 2003 in a prospective cohort study. BV was diagnosed by Gram stain, TV by wet mount, and YV by symptoms with microscopically visible hyphae or positive culture. Trends were assessed using Poisson models., Results: At baseline, BV was present in 42.8% and 47.0% of HIV-infected and uninfected women (P = 0.21), TV in 6.1% and 7.8% (P = 0.17), and YV in 10.0% and 3.8% (P < 0.001). Over time, rates of BV and TV decreased significantly in both groups, whereas rates of YV declined only among HIV-infected women. Risk of BV was not associated with HIV status, whereas HIV-infected women had a lower risk of TV. Highly active antiretroviral therapy (HAART) use was associated with decreased risk of all 3 infections., Conclusions: : Declines in BV, TV, and YV represent decreased morbidity for HIV-infected women and, potentially, decreased risk of transmission of HIV, because each has been associated with increased genital detection of HIV.

Objective: Despite the low mortality and morbidity of major obstetric and gynecologic surgeries (including hysterectomy and cesarean delivery), women undergoing these procedures occasionally suffer from intractable postoperative suprapubic and groin pain. We present seven patients whose intractable pain lasted longer than 6 months and was not due to gynecologic disease or other obvious pathology., Methods: Neuromas of the ilioinguinal, iliohypogastric, and/or genitofemoral nerves were suspected clinically and confirmed intraoperatively., Results: After neuroma resection, all patients reported complete and durable pain relief., Conclusion: Intractable pain after obstetric or gynecologic surgery can be due to neuroma formation, and resection is therapeutic. We suggest an algorithm for the management of women with chronic intractable suprapubic or groin pain after major obstetric and gynecologic surgery., Level of Evidence: II-3.

Objective: To estimate the risk of and risk factors for progression among human immunodeficiency virus (HIV)-seropositive women with abnormal cervical cytology but negative colposcopy., Methods: In a prospective cohort study, 391 HIV-seropositive and 103 seronegative women with cervical cytology read as atypical squamous cells (ASC) or low-grade squamous intraepithelial lesion (LSIL) but negative colposcopy were followed up for a mean of 4.0 years with cytology at 6-month intervals. Colposcopy was prescribed for any epithelial abnormality., Results: Progression to CIN2, CIN3, high-grade SIL/severe dysplasia, or cancer occurred in 47 (12%) HIV-seropositive women and 4 (4%) HIV-seronegative women (P = .02). Progression to CIN1 was seen in an additional 12 HIV-seropositive women and 1 seronegative woman. In multivariate analysis, high-risk but not low-risk HPV detection (hazard ratio [HR] 2.46-95% confidence interval [CI] 1.18-5.12, P = .02 for high risk, HR 1.41, 95% CI 0.62-3.21, P = .42 for low risk), satisfactory colposcopy (HR 2.01, 95% CI 1.11-3.65, P = .02), and non-Hispanic African-American ethnicity (HR 5.08, 95% CI 1.72-14.98, P = .003) were the only factors associated with progression, while HIV serostatus was marginally significant (HR 2.53, 95% CI 0.85-7.50, P = .09)., Conclusion: Human immunodeficiency virus-seropositive women with negative colposcopy after borderline cytology face a higher risk of progression than seronegative women, but the absolute risk is low and becomes nonsignificant after controlling for HPV risk type, ethnicity, and colposcopic findings. Observation is appropriate., Level of Evidence: II-2.

Background: Whether the natural history of human papillomavirus (HPV) infection is affected by bacterial vaginosis (BV) or Trichomonas vaginalis (TV) infection has not been adequately investigated in prospective studies., Methods: Human immunodeficiency virus 1 (HIV-1)-infected (n=1763) and high-risk HIV-1-uninfected (n=493) women were assessed semiannually for BV (by Nugent's criteria), TV infection (by wet mount), type-specific HPV (by polymerase chain reaction with MY09/MY11/HMB01 HPV primers), and squamous intraepithelial lesions (SIL) (by cytological examination). Sexual history was obtained from patient report at each visit. Risk factors for prevalent and incident HPV infection and SIL were evaluated by use of multivariate models., Results: BV was associated with both prevalent and incident HPV infection but not with duration of HPV infection or incidence of SIL. TV infection was associated with incident HPV infection and with decreased duration and lower prevalence of HPV infection. TV infection had no association with development of SIL. Effects of BV and TV infection were similar in HIV-1-infected and high-risk HIV-1-uninfected women. HIV-1 infection and low CD4(+) lymphocyte count were strongly associated with HPV infection and development of SIL., Conclusions: BV and TV infection may increase the risk of acquisition (or reactivation) of HPV infection, as is consistent with hypotheses that the local cervicovaginal milieu plays a role in susceptibility to HPV infection. The finding that BV did not affect persistence of HPV infection and that TV infection may shorten the duration of HPV infection helps explain the lack of effect that BV and TV infection have on development of SIL.

Objective: We sought to estimate rates of progression and regression of grade 1 cervical intraepithelial neoplasia (CIN 1) among women with human immunodeficiency virus (HIV)., Methods: In a multicenter prospective cohort study, HIV-seropositive and HIV-seronegative women were evaluated colposcopically after receiving an abnormal cytology test result between November 1994 and September 2002. Women with CIN 1 were included, except those who had undergone hysterectomy, cervical therapy, or had CIN 2-3 or cervical cancer. Those women who were included were followed cytologically twice yearly, with colposcopy repeated for atypia or worse., Results: We followed 223 women with CIN 1 (202 HIV seropositive and 21 HIV seronegative) for a mean of 3.3 person-years. Progression occurred in 8 HIV-seropositive women (incidence density, 1.2/100 person-years; 95% confidence interval [CI] 0.5-2.4/100 person-years) and in no HIV seronegative women. Regression occurred in 66 (33%) HIV-seropositive women (13/100 person-years, 95% CI 10-16/100 person-years) versus 14 (67%) seronegative women (32/100 person-years, relative risk 0.40, 95% CI 0.25-0.66; P < .001). In multivariate analysis, regression was associated with human papillomavirus (HPV) detection (hazard ratio [HR] for low risk 0.28, 95% CI 0.13-0.61, P = .001; and for high-risk 0.34, 95% CI 0.20-0.55, P < .001 versus no HPV detected) and Hispanic ethnicity (HR 0.48, 95% CI 0.230.98; P = .04); HIV serostatus was only marginally linked to regression (HR 0.52, 95% CI 0.27-1.03; P = .06), but seropositive women were less likely to regress when analysis was limited to 146 women with HPV detected at CIN 1 diagnosis (HR 0.18, 95% CI 0.05-0.62; P = .006)., Conclusion: Grade 1 cervical intraepithelial neoplasia infrequently progresses in women with HIV. Thus, observation appears safe absent other indications for treatment., Level of Evidence: II-1.

Background: Women infected with human immunodeficiency virus (HIV) have an increased risk of persistent squamous intraepithelial lesions (SILs) of the cervix. We assessed the association between use of highly active antiretroviral therapy (HAART) and regression of SIL in HIV-infected women enrolled in the Women's Interagency HIV Study, a large, multicenter, prospective cohort study., Methods: Of 2059 HIV-infected participants, 312 HIV-infected women had normal cervical cytology at baseline and were subsequently diagnosed during 7 years of follow-up with incident SIL. Pap smears, CD4+ T-cell counts, and information regarding use of HAART were obtained every 6 months. The outcome of interest was lesion regression, defined as two consecutive normal Pap smears 6 months apart. Incidence rates of SIL regression were computed among person-years at risk, both before and after HAART initiation. All statistical tests were two-sided., Results: Of 312 women, 141 had lesions that regressed to normal cytology, with a median time to regression of 2.7 years. Overall, the incidence of regression increased (P(trend) =.002) over time after HAART was introduced. At incident SIL, median CD4+ T-cell counts were lower in women whose lesions did not regress than in women whose lesions regressed (230 versus 336 cells/microL; P<.01). Before HAART was introduced, the rate of lesion regression was 0.0% (95% confidence interval [CI' = 0.0% to 2.4%). After HAART was introduced, the rate was 12.5% (95% CI = 9.9% to 15.1%) and was related to post-HAART CD4+ T-cell counts (P(trend) =.002)., Conclusions: HAART use was associated with increased regression of SIL among HIV-infected women, and among women who used HAART, increased CD4+ T-cell counts were associated with a greater likelihood of regression. However, the majority of cervical lesions among HIV-infected women, even among individuals who used HAART, did not regress to normal.

Objective: The purpose of this study was to determine the incidence and predictors of genital warts and vulvar intraepithelial neoplasia among women with the human immunodeficiency virus., Study Design: This was a multicenter prospective cohort study comprised of women without warts or vulvar intraepithelial neoplasia at baseline who underwent CD4 count, human immunodeficiency virus RNA measurement, examination, Papanicolaou test, and biopsy, as indicated, every 6 months. Human papillomavirus DNA typing was examined at baseline., Results: The incidence of warts among women who were human immunodeficiency virus seronegative was 1.31 versus 5.01 per 100 person-years among women who were seropositive (P < .001). Incidence of vulvar intraepithelial neoplasia among women who were seronegative was 1.31 versus 4.67 per 100 person-years among women who were seropositive (P < .001). In multivariable analysis, warts were associated with highly active antiretroviral therapy (relative hazard, 0.76), CD4 count (relative hazard, 0.91/100 cell/cm(2) increase), acquired immunodeficiency syndrome (relative hazard, 1.25), abnormal Papanicolaou test results (relative hazard, 2.18), high- or medium-risk human papillomavirus types (relative hazard, 1.91), low-risk human papillomavirus types (relative hazard, 1.48), smoking (relative hazard, 1.43), having 1 child (relative hazard, 1.54), and age (relative hazard, 0.74/10 years). Vulvar intraepithelial neoplasia was linked to highly active antiretroviral therapy (relative hazard, 0.65), CD4 count (relative hazard, 0.92), abnormal Papanicolaou test results (relative hazard, 16.03), high- or medium-risk human papillomavirus types (relative hazard, 1.37), and age (relative hazard, 0.85/10 years)., Conclusion: Warts and vulvar intraepithelial neoplasia are common among women with human immunodeficiency virus. Highly active antiretroviral therapy decreases their incidence.

Objective: To determine incidence, progression, and regression rates for abnormal cervical cytology and their correlates among women with HIV., Methods: In a multicenter prospective cohort study conducted October 1, 1994, through September 30, 1999 at university, public, and private medical centers and clinics, 1639 HIV-seropositive and 452 seronegative women were evaluated every 6 months for up to 5 years using history, cervical cytology, T-cell subsets, and quantitative plasma HIV RNA. Human papillomavirus (HPV) typing at baseline was determined by polymerase chain reaction. Cytology was read using the Bethesda system, with any smear showing at least atypia considered abnormal. Poisson regression identified factors associated with incident cytologic abnormalities whereas logistic regression identified those associated with progression and regression after an abnormality., Results: At least one abnormal smear was found during all of follow-up among 73.0% of HIV-seropositive patients and 42.3% of seronegatives (p <.001). Only 5.9% of seropositives ever developed high-grade lesions, and the proportion with high-grade findings did not rise over time. Incidence of atypical squamous cells of uncertain significance (ASCUS) or more severe lesions among HIV-seropositive patients and seronegative patients was 26.4 and 11.0/100 woman-years (rate ratio [RR], 2.4; 95% confidence interval [CI], 1.9-3.0), whereas that of at least low-grade squamous intraepithelial lesions (SIL) was 8.9 and 2.2/100 (RR, 4.0; CI, 2.6-6.1). HIV status, detection of the presence of human papillomavirus (HPV), CD4 lymphocyte count, and HIV RNA level predicted incidence of abnormal cytology (p <.05); HPV detection and HIV RNA level predicted progression (p <.01); and HPV detection, CD4 lymphocyte count, and HIV RNA level predicted regression (p <.001). Rates of incidence, progression, and regression of abnormal cytology did not differ between HIV seronegative women and seropositive women with CD4 lymphocyte counts >200/mm(3) and HIV RNA levels <4000/ml of similar HPV status., Conclusions: Although HIV infected women were at high risk for abnormal cytology, high-grade changes were uncommon. HIV status, HPV detection, CD4 lymphocyte count, and HIV RNA level predicted the incidence of cervical cytologic abnormalities. Progression was significantly increased only among the most immunosuppressed women, while regression was significantly reduced in all HIV seropositive women except those with the best controlled HIV disease.

This study reports that quiescent herpes simplex virus (HSV) type 1 genomes, persisting in long-term infected nerve growth factor (NGF) differentiated PC12 cells, were not stimulated by superinfection with a HSV-1. We have previously shown that HSV-1 can establish long term, quiescent infections in NGF differentiated PC12 cells. To determine if virion associated factors or virus induced gene products could trans-activate the quiescent viral genomes, long term infected PC12 cell cultures were superinfected at a high moi (moi of 20) with a recombinant HSV 17alpha47/lacZ that contains the lacZ gene within the alpha 47 locus. Progeny virus and gene expression from the resident 'quiescent' viral genomes were not detected following superinfection with recombinant 17alpha47/lacZ. The failure to stimulate the quiescent genome appears to be related to the inability of the super infecting virus to induce any gene expression from its own genome following entry into the long term NGF treated PC12 cells. Interestingly, both primary and superinfecting viruses could be stimulated from the quiescently infected cultures following cocultivation with inducer cells. These data suggest that (i) HSV genomes in quiescently infected PC12 cells are unable to be stimulated by incoming virion associated factors and (ii) NGF differentiated PC12 cells maintained in tissue culture for longer than 3 weeks became completely refractory to viral gene expression. The possibilities that these results are reflective of populations of neural cells, in vivo in mouse central nervous system, which are completely refractory to virus gene expression, yet accommodating to the maintenance of viral genomes and thus favor 'latency', are discussed.

Objective: To evaluate lung transplantation for treatment of surfactant protein B (SP-B) deficiency., Study Design: We compared surfactant composition and function from pretransplantation and posttransplantation samples of bronchoalveolar lavage fluid, somatic and lung growth, neurodevelopmental progress, pulmonary function, and pulmonary immunohistology in 3 infants with SP-B deficiency who underwent bilateral lung transplantation at 2 months of age and 3 infants who underwent lung transplantation for other reasons., Results: Two years after transplantation, the 2 surviving infants with SP-B deficiency exhibited comparable somatic growth and cognitive development to the comparison infants. All infants had delays in gross motor development that improved with time. Both groups have exhibited normal gas exchange, lung growth, and pulmonary function. The SP-B-deficient infants have also exhibited normal SP-B expression and pulmonary surfactant function after lung transplantation. In two SP-B-deficient infants antibody to SP-B developed. No pathologic consequences of this antibody were identified., Conclusions: Apart from the development of anti-SP-B antibody, the outcomes for SP-B-deficient infants after lung transplantation are similar to those of infants who undergo lung transplantation for other reasons. Lung transplantation offers a successful interim therapy until gene replacement for this disease is available.

The 5' flanking region of the Clara cell secretory protein (CCSP) gene contains two cis-acting elements which bind hepatocyte nuclear factor (HNF)-3 alpha and HNF-3 beta in vitro. To determine the role of these proteins in mediating CCSP gene expression in the bronchiolar epithelium, chimeric CCSP-reporter gene constructs containing various regions of the CCSP 5' flanking region were co-transfected into H-441 cells with HNF-3 alpha or HNF-3 beta expression plasmids. These studies indicate that each of these transcription factors positively regulates CCSP gene expression and revealed that CCSP region I (-132 to -76) is sufficient to mediate this effect. Gel-mobility-shift assays with oligonucleotides corresponding to CCSP region I, nuclear extract from bronchiolar epithelial cells and HNF-3-specific antibodies indicate that HNF-3 alpha and HNF-3 beta are the only proteins in bronchiolar epithelial cells which directly interact with this region. Consistent with these observations, HNF-3 alpha and HNF-3 beta transcripts were found to be enriched in this cell population and in situ hybridization of adult lung revealed HNF-3 gene expression in non-ciliated bronchiolar epithelial cells expressing the CCSP gene. Finally, experiments with CCSP region I and a heterologous promoter indicate that this region acts in a promoter-specific context, suggesting that additional factors interacting via the minimal CCSP promoter region are essential in determining the effects of HNF-3 on cell-specific CCSP gene expression in the bronchiolar epithelium.

An infant with a family history of congenital alveolar proteinosis associated with surfactant protein B (SP-B) deficiency was identified when SP-B was not detected in amniotic fluid obtained at 37, 38, and 40 weeks of gestation. Surfactant replacement with commercially available preparations that contained SP-B was begun soon after delivery. Progressive respiratory failure developed despite continued surfactant replacement, corticosteroid therapy, and extracorporeal membrane oxygenation. The infant died at 54 days of age while awaiting lung transplantation. Surfactant extracted from amniotic fluid, bronchoalveolar lavage fluid, and lung tissue had no phosphatidylglycerol; surface tension was 24 dynes/cm (normal, < 10 dynes/cm) and did not decrease with in vitro addition of exogenous SP-B. Pulmonary vascular permeability measured with positron emission tomography was twice normal. At autopsy the alveolar proteinosis pattern was less prominent than that seen in affected siblings. Immunoreactivity of SP-B was absent in type II cells, but numerous foreign body granulomas with central immunoreactivity for SP-B and surfactant protein C were present. We conclude that exogenous surfactant replacement did not normalize surfactant composition, activity, or pulmonary vascular permeability. These findings suggest that endogenous SP-B synthesis is necessary for mature surfactant metabolism and function.

Surfactant protein D (SP-D) is a collagenous, surfactant-associated, carbohydrate-binding protein that is synthesized by alveolar type II epithelial cells. To further characterize SP-D, we isolated RNA from adult rat lungs and rat type II cells and translated mRNAs in vitro. [35S]methionine-labeled translation products were precipitated with antibodies to rat SP-D, resolved by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and visualized by fluorography. Immune precipitates of translation reactions for rat lung or rat type II cells demonstrated a single collagenous polypeptide (39.3 kDa) that was smaller than surfactant-associated SP-D (43 kDa, reduced) but larger than the mature secreted form of rat SP-A. This component was not identified in translation reactions of rat liver, gut, brain, mammary gland, or rat L2 cell RNA. There was a fivefold enrichment of SP-D mRNA in freshly isolated type II cells relative to lung; however, the levels of translatable SP-D mRNA decreased rapidly during the first 24 h of cell culture. The SP-D translation product migrated faster than the major cellular form of SP-D but approximately 1 kDa slower than cellular SP-D synthesized in the presence of 2,2'-dipyridyl plus tunicamycin. Translation in the presence of canine pancreatic microsomes gave a single glycosylated, endoglycosidase F-sensitive form (40.6 kDa) and demonstrated cleavage of a small signal peptide. These results indicate that SP-D is a secretory product of differentiated type II epithelial cells and that SP-D is secreted in a mature form that does not undergo further proteolytic processing in vivo.

Human lymphoblastoid cell lines evolve in vitro by the emergence of successive waves of clones which are often chromosomally marked. This offers the opportunity to compare tissue samples of the same genetic origin but differing in certain defined parts of the karyotype. Using selected sets of lines in which the members of genetically matched pairs differed in the number of copies of 8p or of 12p, levels of GSR and LDH B respectively have been shown to correlate with the specific chromosome aberrations, supporting existing data on the regional assignment of these two structural loci. This approach represents a useful addition to established methods for human gene mapping.

Conventional cleavage of linked polypeptide chains by heating in SDS can so alter molecular structure as to interfere with antibody binding, on which both immunoprecipitation and 'western blotting' depend. As an alternative, gentle treatment with acid at room temperature or at 0 degrees C was effective in separating the alpha and beta chains of human MHC Class II glycoprotein dimers and proved superior in terms of preservation of at least one labile epitope on the beta chain.

CP4 is a collagenous glycoprotein (43 kDa, reduced) synthesized by rat type II pulmonary epithelial cells in primary culture (Persson et al., 1988). In order to better characterize this protein, CP4 was isolated from rat bronchoalveolar lavage and EDTA extracts of lung surfactant by adsorption to barium sulfate and elution with sodium citrate followed by reverse-phase HPLC. Amino acid analysis of purified CP4 demonstrated 4-hydroxyproline (Hyp), hydroxylysine (Hyl), and acid-labile components coeluting with Hyl glycosides. In addition, gas-phase amino-terminal microsequencing of two CP4 CNBr peptides demonstrated nonoverlapping collagenous sequences comprised of nine and six Gly-X-Y triplets, containing a total of four residues of Hyp and two of Hyl. There was less than 50% sequence homology of these peptides with the cDNA-derived sequence of the collagenous domain of rat SP-A. Two-dimensional IEF/SDS-PAGE resolved the protein into a charge train of basic isoforms (pI approximately 6-8), similar to those of newly synthesized CP4 and the class D surfactant proteins (Phelps & Taeusch, 1985). Gel filtration of nondenatured CP4 on 4% agarose showed a high apparent molecular mass complex comprised of disulfide-bonded trimers of the 43-kDa subunits. Antibodies to purified lavage CP4 showed specific binding to newly synthesized and surfactant-associated CP4. We propose that CP4 be designated "surfactant protein D" (SP-D) in accordance with an accepted nomenclature for surfactant-associated proteins.

Ia antigens were shown to be present in the cell almost exclusively as mature alpha beta dimers which split into separate alpha and beta chains after boiling in SDS. In contrast metabolically labelling the cells with [35S]methionine resulted in only free alpha and beta chains being labelled. It is concluded that this widely used type of labelling, although useful for studying intermediate synthesis, should not be used for labelling mature cell surface molecules.

Monoclonal antibodies directed against human Ia alpha- and beta-subunit chains have been used as probes to detect polypeptides carrying recognized antigenic determinants or epitopes following two-dimensional PAGE separation. Approximately 4 sets of components differing in isoelectric point but not in molecular weight are recognized by each antibody. The anti-alpha-chain, McAb, reacts weakly with spots designated as epsilon but neither antibody recognizes Im, Ii or delta determinants under the conditions tested.

Type II pneumocytes secrete pulmonary surfactant and are known to synthesize SP-35, a collagenous surfactant-associated protein. Freshly isolated type II cells also synthesize other bacterial collagenase-sensitive and hydroxyproline-containing proteins, including a glycoprotein designated CP4. CP4 was isolated from rat pneumocyte culture medium by immune precipitation with polyclonal antibodies to rat surfactant proteins or by DEAE chromatography and reverse-phase or gel permeation HPLC. CP4 did not cross-react with polyclonal antibodies to SP-35 and was completely resolved from SP-35 by SDS-PAGE (Mr 43K reduced) or isoelectric focusing. Unlike SP-35, which consists of acidic isoforms assembled as disulfide-bonded dimers and multimers, CP4 was secreted as basic isoforms assembled as disulfide-bonded trimers. Differences in primary structure were demonstrated by CNBr and V8 protease peptide mapping. The secretion of both proteins was inhibited by 2,2'-dipyridyl, an inhibitor of posttranslational prolyl and lysyl hydroxylation and collagen triple helix formation. CP4 was isolated from EDTA extracts of rat surfactant. These studies provide evidence for the heterogeneity of pneumocyte-derived collagenous surfactant-associated proteins.

Four mouse monoclonal antibodies to human B cell surface determinants previously described as being directed against Ia like (MHC class II) antigens, have been shown to precipitate Ia alpha and beta chains. Electrophoretic transfer experiments showed one antibody to be directed against Ia alpha chains and two others to be against Ia beta chains. The antibodies were then used to analyse a range of cell types and a large number of lymphoblastoid and lymphoma cell lines. Ia antigens could not be detected on peripheral blood T cells, cord endothelium or T cell lines but their presence was confirmed on activated T cells and peripheral blood non-T cells. There was both qualitative and quantitative variation of Ia like antigen expression on B cell lines, including an apparent genetic polymorphism in alpha chain structure unrelated to DR allotypes and a single instance of a beta chain of abnormally high molecular weight.