clinicaltrials.gov Identifier: NCT02139930.

Matched-pair design is often used in clinical trials to increase the efficiency of treatment comparison. We consider the problem of equivalence test with a relative risk endpoint in matched-pair studies with binary outcomes, and develop several score and Wald-type statistics for testing a hypothesis of non-unity relative risk. Examples from an assessment of HIV screening test and a cross-over clinical trial of soft contact lenses are used to illustrate the proposed methods. Through simulations we compare the empirical performance of these tests with the test proposed by Lachenbruch and Lynch. We show that a score test based on a reparameterized multinomial model by Tango performs best in the sense that the test satisfactorily controls the type I error rate and its empirical type I error rates are generally much closer to the prespecified nominal significance level than those of the other tests.

SUMMARY During pancreatic development, proliferating pancreatic progenitors activate the proendocrine transcription factor neurogenin 3 (NEUROG3), exit the cell cycle, and differentiate into islet cells. The mechanisms that direct robust NEUROG3 expression within a subset of progenitor cells control the size of the endocrine population. Here we demonstrate that NEUROG3 is phosphorylated within the nucleus on serine 183, which catalyzes its hyperphosphorylation and proteosomal degradation. During progression through the progenitor cell cycle, NEUROG3 phosphorylation is driven by the actions of cyclin-dependent kinases 2 and 4/6 at G1/S cell-cycle checkpoint. Using models of mouse and human pancreas development, we show that lengthening of the G1 phase of the pancreatic progenitor cell cycle is essential for proper induction of NEUROG3 and initiation of endocrine cell differentiation. In sum, these studies demonstrate that progenitor cell-cycle G1 lengthening, through its actions on stabilization of NEUROG3, is an essential variable in normal endocrine cell genesis.

Tumors are believed to emerge only when immune surveillance fails. We wished to ascertain whether the failure to inherit putative protective alleles of HLA class II genes is linked to the development of breast cancer. We molecularly typed HLA DPB1, DQB1, DRB1, and DRB3 alleles in 176 Caucasian women diagnosed with early-onset breast cancer and in 215 ethnically matched controls. HLA DQB*03032 was identified in 7% of controls but in no patients with early-onset breast cancer (P ‫؍‬ 0.0001). HLA DRB1*11 alleles were also significantly overrepresented (P < 0.0001) in controls (16.3%) as compared with patients with early-onset breast cancer (3.5%). HLA DQB*03032 and HLA DRB1*11 alleles may have a protective role in human breast cancer.A lthough it is likely that genetic susceptibility plays a role in the development of most human cancers, evidence supporting this view hitherto has been obtained in only a small fraction of patients who typically carry germ-line mutations in tumor suppressor genes. In addition to the widely recognized role of acquired alterations in oncogenes and tumor suppressor genes, considerable evidence exists to suggest that the immune system might play a protective role in tumorigenesis. Although immune surveillance is believed to be involved in the elimination of tumors (1, 2), immunotherapeutic approaches to human cancer by and large have proved unsuccessful.T cell responses are dependent on the inheritance of specific alleles of the highly polymorphic HLA class I and class II genes. Although weak associations of specific HLA alleles with tumors of viral origin have been described (3-10), the relevance of MHC polymorphisms to the broader category of spontaneous nonviral human tumors remains to be established. Somatic alterations in many tumors can contribute to the down-regulation of HLA class I gene expression in tumor cells (11). These alterations potentially could contribute to immune evasion and might represent a discrete event in the multistep paradigm of tumorigenesis. Although HLA class I genes are expressed in all cells, immune responses also require the presentation of antigenic peptides to T cells by HLA class II molecules. These heterodimers primarily are expressed by professional antigen-presenting cells such as macrophages, dendritic cells, and B lymphocytes. Somatic alterations in tumor cells cannot inf luence the expression of HLA class II genes in dendritic cells and other professional antigen-presenting cells. If, indeed, immune surveillance is important during tumorigenesis, certain individuals who inherit specific alleles of the highly polymorphic HLA class II DPB, DQB, or DRB genes might be resistant to specific types of cancers.We reasoned that a detailed molecular analysis of HLA DPB, DQB, and DRB alleles in patients with breast cancer and ethnically matched controls might provide information on the potential existence of alleles that could confer susceptibility or resistance to this human cancer. Women with early-onset breast cancer (diagnosed at or before the age of 40...

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Introduction Electronic cigarettes (e-cigarettes) have the potential to significantly reduce exposure to harmful constituents associated with cigarette smoking when smokers completely substitute cigarettes with e-cigarettes. This study examined patterns of e-cigarette and cigarette use, and extent of toxicant exposure, if smokers were instructed and incentivized to completely switch to e-cigarettes compared to instructions to use the product ad libitum. Aims and Methods US adult daily smokers (n = 264; 49.2% female; Mage = 47.0), uninterested in quitting smoking immediately, were recruited from Minneapolis, MN, Columbus, OH, and Buffalo, NY. Participants were randomized to 8 weeks of instructions for (1) ad libitum use of e-cigarettes (AD-E), (2) complete substitution of cigarettes with e-cigarettes (CS-E), (3) complete substitution of cigarettes with nicotine gum or lozenge (CS-NRT), or (4) continue smoking of usual brand cigarettes (UB). Participants were incentivized for protocol compliance, including complete switching in the CS-E and CS-NRT groups. Outcome variables were cigarette smoking rate and tobacco-related biomarkers of exposure. Results Smokers in the CS-E and CS-NRT groups showed lower rates of smoking and lower exposure to carbon monoxide, tobacco carcinogens, and other toxicants than smokers in the AD-E group. In general, no significant differences were observed between CS-E versus CS-NRT or between AD-E versus UB for most biomarkers. Significantly higher 7-day point prevalence smoke-free rates were observed for CS-E versus CS-NRT. Conclusions Smokers instructed and incentivized to completely switch to e-cigarettes resulted in lower smoking rates and greater reductions in exposures to harmful chemicals than smokers instructed to use the product ad libitum. Implications Smokers instructed to completely substitute e-cigarettes for cigarettes displayed significantly lower levels of smoking and biomarkers of exposure to carcinogens and toxicants, compared to smokers instructed to use e-cigarettes ad libitum and similar levels as smokers instructed to completely substitute with nicotine replacement therapies. Furthermore, a higher rate of complete switching was achieved with e-cigarettes versus nicotine replacement therapies. Approaches to maximize complete substitution with e-cigarettes are an important area for future research.

The human umbilical cord perivascular cells (HUCPVCs) have been considered as an alternative source of mesenchymal progenitors for cell based regenerative medicine. However, the biological properties of these cells remain to be well characterized. In the present study, HUCPVCs were isolated and sorted by CD146+ pericyte marker. The purified CD146+ HUCPVCs were induced to differentiate efficiently into osteoblast, chondrocyte and adipocyte lineages in vitro. Six weeks following subcutaneous transplantation of CD146+ HUCPVCs-Gelfoam-alginate 3D complexes in severe combined immunodeficiency (SCID) mice, newly formed bone matrix with embedded osteocytes of donor origin was observed. The functional engraftment of CD146+ HUCPVCs in the new bone regenerates was further confirmed in a critical-sized bone defect model in SCID mice. Hypoxic conditions suppressed osteogenic differentiation while increased cell proliferation and colony-forming efficiency of CD146+ HUCPVCs as compared to that under normoxic conditions. Re-oxygenation restored the multi-differentiation potential of the CD146+ HUCPVCs. Western blot analysis revealed an upregulation of HIF-1α, HIF-2α, and OCT-4 protein expression in CD146+ HUCPVCs under hypoxia, while there was no remarkable change in SOX2 and NANOG expression. The gene expression profiles of stem cell transcription factors between cells treated by normoxia and hypoxic conditions were compared by PCR array analysis. Intriguingly, PPAR-γ was dramatically downregulated (20-fold) in mRNA expression under hypoxia, and was revealed to possess a putative binding site in the Hif-2α gene promoter region. Chromatin immunoprecipitation assays confirmed the binding of PPAR-γ protein to the Hif-2α promoter and the binding was suppressed by hypoxia treatment. Luciferase reporter assay showed that the Hif-2α promoter activity was suppressed by PPAR expression. Thus, PPAR-γ may involve in the regulation of HIF-2α for stemness maintenance and promoting the expansion of CD146+ HUCPVCs in response to hypoxia. CD146+ HUCPVCs may serve as a potential autologous cell source for bone regeneration.

The growth arrest specific 1 (gas1) gene is highly expressed in quiescent mammalian cells (Schneider et al., 1988, Cell 54, 787-793). Overexpression of gas1 in normal and some cancer cell lines could inhibit G(0)/G(1) transition. Presently, we have examined the functions of this gene in the developing mouse embryo. The spatial-temporal expression patterns for gas1 were established in 8.5- to 14.5-day-old embryos by immunohistochemical staining and in situ hybridization. Gas1 was found heterogeneously expressed in most organ systems including the brain, heart, kidney, limb, lung, and gonad. The antiproliferative effects of gas1 on 10.5 and 12.5 day limb cells were investigated by flow cytometry. In 10.5 day limbs cells, gas1 overexpression could not prevent G(0)/G(1) progression. It was determined that gas1 could only induce growth arrest if p53 was also coexpressed. In contrast, gas1 overexpression alone was able to induce growth arrest in 12.5 day limb cells. We also examined the cell cycle profile of gas1-expressing and nonexpressing cells by immunochemistry and flow cytometry. For 10.5 day Gas1-expressing heart and limb cells, we did not find these cells preferentially distributed at G0/G1, as compared with Gas1-negative cells. However, in the 12.5 day heart and limb, we did find significantly more Gas1-expressing cells distributed at G0/G1 phase than Gas1-negative cells. These results implied that Gas1 alone, during the early stages of development, could not inhibit cell growth. This inhibition was only established when the embryo grew older. We have overexpressed gas1 in subconfluent embryonic limb cells to determine the ability of gas1 to cross-talk with various response elements of important transduction pathways. Specifically, we have examined the interaction of gas1 with Ap-1, NFkappaB, and c-myc responsive elements tagged with a SEAP reporter. In 10.5 day limb cells, gas1 overexpression had little effect on Ap-1, NFkappaB, and c-myc activities. In contrast, gas1 overexpression in 12.5 day limb cells enhanced AP-1 response while it inhibited NFkappaB and c-myc activities. These responses were directly associated with the ability of gas1 to induce growth arrest in embryonic limb cells. In the 12.5 day hindlimb, gas1 was found strongly expressed in the interdigital tissues. We overexpressed gas1 in these tissues and discovered that it promoted interdigital cell death. Our in situ hybridization studies of limb sections and micromass cultures revealed that, during the early stages of chondrogenesis, only cells surrounding the chondrogenic condensations expressed gas1. The gene was only expressed by chondrocytes after the cartilage started to differentiate. To understand the function of gas1 in chondrogenesis, we overexpressed the gene in limb micromass cultures. It was found that cells overexpressing gas1/GFP could not participate in cartilage formation, unlike cells that just express the GFP reporter. We speculated that the reason gas1 was expressed outside the chondrogenic nodules was to restrict cells fro...