Since human papillomavirus (HPV) infection is strongly associated with cervical neoplasia and tumor hypoxia has prognostic significance in human cervical carcinomas, we examined the relationship ...between hypoxia and apoptosis in human cervical epithelial cells expressing high-risk HPV type 16 oncoproteins. In vitro, hypoxia stimulated both p53 induction and apoptosis in primary cervical epithelial cells infected with the HPV E6 and E7 genes but not in cervical fibroblasts infected with E6 and E7. Furthermore, cell lines derived from HPV-associated human cervical squamous cell carcinomas were substantially less sensitive to apoptosis induced by hypoxia, indicating that these cell lines have acquired additional genetic alterations that reduced their apoptotic sensitivity. Although the process of long-term cell culturing resulted in selection for subpopulations of HPV oncoprotein-expressing cervical epithelial cells with diminished apoptotic potential, the exposure of cells to hypoxia greatly accelerated the selection process. These results provide evidence for the role of hypoxia-mediated selection of cells with diminished apoptotic potential in the progression of human tumors and can in part explain why cervical tumors that possess low pO2 values are more aggressive.
Zika virus (ZIKV) has gained worldwide attention since it emerged, and a global effort is underway to understand the correlates of protection and develop diagnostics to identify rates of infection. ...As new therapeutics and vaccine approaches are evaluated in clinical trials, additional effort is focused on identifying the adaptive immune correlates of protection against ZIKV disease. To aid in this endeavor we have begun to dissect the role of CD4+T cells in the protection against neuroinvasive ZIKV disease. We have identified an important role for CD4+T cells in protection, demonstrating that in the absence of CD4+T cells mice have more severe neurological sequela and significant increases in viral titers in the central nervous system (CNS). The transfer of CD4+T cells from ZIKV immune mice protect type I interferon receptor deficient animals from a lethal challenge; showing that the CD4+T cell response is necessary and sufficient for control of ZIKV disease. Using a peptide library spanning the complete ZIKV polyprotein, we identified both ZIKV-encoded CD4+T cell epitopes that initiate immune responses, and ZIKV specific CD4+T cell receptors that recognize these epitopes. Within the ZIKV antigen-specific TCRβ repertoire, we uncovered a high degree of diversity both in response to a single epitope and among different mice responding to a CD4+T cell epitope. Overall this study identifies a novel role for polyfunctional and polyclonal CD4+T cells in providing protection against ZIKV infection and highlights the need for vaccines to develop robust CD4+T cell responses to prevent ZIKV neuroinvasion and limit replication within the CNS.
HPV-16 E6 and E7 genes are required to efficiently immortalize a broad spectrum of cell types including cervical keratinocytes. Therefore, the E6/E7 genes can be considered relevant targets for ...anti-cancer therapy. We produced several engineered hairpin (HP) ribozymes to specifically disrupt HPV-16 E6/E7 mRNA. After extensive biochemical characterization, one anti-E6 HP ribozyme (R434) was selected for in vivo testing because of its superior catalytic capabilities. When expressed in cis, R434 efficiently inhibited E6 in vitro translation. Cis-expression of the HP ribozyme with HPV-16 E6/E7 genes in normal human keratinocytes reduced the growth rate and prevented immortalization. RNA analysis by reverse transcription-PCR showed that E6/E7 transcripts were cleaved in post-transfected cells and virtually were eliminated after long term expression. Of interest, an inactive version of the HP also was able to significantly affect the immortalizing ability of E6/E7, probably through passive hybridization. The combination of passive and cleaving anti-sense RNA therefore is established as an effective inhibitor of HPV-16 E6/E7 immortalization.
Extreme weather events, such as hurricanes, can be ecologically devastating and cause widespread mortality. Recent studies in Anolis lizards report hurricane‐induced phenotypic shifts and selection ...favouring morphological variation related to clinging performance. Although it is difficult to observe organismal responses during extreme events in nature, we can experimentally simulate the high‐speed winds associated with hurricanes to evaluate the putative mechanism underlying observed patterns of natural selection.
In this study, we used two laboratory experiments to better understand the clinging performance and behaviour of Anolis lizards when experiencing hurricane‐force winds. We assessed the physical ability of lizards when using the combined function of their claws, limbs, toepads and other traits to resist forces pulling them off a perch. We also evaluated the combination of this physical clinging ability of lizards and their behavioural responses to avoid being blown off a perch during high winds. We assessed behaviour that could decrease exposure of lizards to wind and increase their clinging ability.
Clinging force measurements revealed variation in performance among species and substrates not reflected in clinging times for lizards experiencing hurricane‐force winds, revealing the importance of behaviour when experiencing high winds. The most arboreal species (A. carolinensis) had substantially longer clinging times on rough substrates compared with the other species, presumably due to its larger toepads for increased clinging as well as its shorter limbs that reduced drag.
Under high‐speed winds, lizards commonly shifted to the more protected leeward side of dowels, especially on broad and rough substrates, presumably to reduce exposure. This reveals how behaviour can mediate factors influencing clinging ability during hurricanes and, in conjunction with ecologically relevant variation in morphology and substrate properties, contribute to clinging performance.
Our experiments reveal that behaviour strongly influences clinging performance during high winds beyond that predicted by physical traits alone. Thus, microhabitat selection of perches and the position of a lizard on its perch during a hurricane will likely have important consequences for clinging performance. This may alter how selection acts on morphological traits and influence the susceptibility of different species to these extreme weather events.
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Read the free Plain Language Summary for this article on the Journal blog.
Chronic inflammation caused by infection with Helicobacter pylori and autoimmune gastritis increases an individual’s risk of developing gastric cancer. More than 90% of gastric cancers are ...adenocarcinomas, which originate from epithelial cells in the chronically inflamed gastric mucosa. However, only a small subset of chronic gastritis patients develops gastric cancer, implying a role for genetic and environmental factors in cancer development. A number of DNA polymorphisms that increase gastric cancer risk have mapped to genes encoding cytokines. Many different cytokines secreted by immune cells and epithelial cells during chronic gastritis have been identified, but a better understanding of how cytokines regulate the severity of gastritis, epithelial cell changes, and neoplastic transformation is needed. This review summarizes studies in both human and mouse models, describing a number of different findings that implicate various cytokines in regulating the development of gastric cancer.
The human involucrin gene, which encodes a precursor of the keratinocyte cornified layer, is strongly expressed in response to differentiation stimuli. Earlier studies suggested that YY1 and ...components of the AP-1 family might participate in the silencing of involucrin in proliferating keratinocytes. This study shows that overexpression of either YY1 or c-Jun represses transcription of the human involucrin gene in multiplying keratinocytes. Transient overexpression and site-directed mutagenesis experiments of the involucrin 5'-non-coding region (5'-NCR) confirmed that YY1 and c-Jun repress involucrin transcription. This repression involves the distal zinc fingers of YY1 protein and the DNA binding and leucine zipper domains of c-Jun. The results with protein pull-down experiments are consistent with the hypothesis that interaction of YY1 with c-Jun is an important mechanism for involucrin repression. Cotransfection of YY1 modified the stimulatory function of mutant c-Jun proteins independently of their DNA binding capacity suggesting that interactions may be more complex in vivo. Additionally, c-Jun protein levels are affected by differentiation stimuli indicating the importance of c-Jun in the YY1 repression pathway. Thus YY1 and c-Jun have an important role in epidermal differentiation by negatively regulating the human involucrin gene.
Cervical cancer is the second most common malignancy in women worldwide. Two HPV strains, HPV-16 and 18, occur in the 70% of untreated cancers. Expression of viral oncogenes E6 and E7 disrupt the ...cell cycle by interfering with p53 and p107(Rb). It is known that HPV infection is necessary but insufficient to cause malignancy. Furthermore, persistence of HPV-16 or 18 in women does not necessarily result in cancer. Persistence indicates the importance of other factors for malignant conversion of high-grade HPV infection. The multi-step cervical carcinogenesis process is amendable to molecular therapeutics such as therapeutic nucleic acids (TNAs). TNA-based therapies for cervical carcinoma include ribozymes, antisense oligonucleotides (AS-ODNs) and small interfering RNAs (siRNAs). In vitro experiments with TNAs successfully inhibited E6/E7 expression and caused induction of apoptosis and/or senescence in cervical carcinoma cells. Early ribozyme and AS-ODN approaches showed promise as therapeutic moieties for cervical cancer. Despite the very high in vitro efficiency of siRNA-based therapies they present the same issues that burdened clinical development of ribozymes and AS-ODNs. These issues include intracellular target accessibility, specificity and delivery. Ribozymes are useful for functional genomic studies including diagnosis. Moreover, AS-ODNs appear better suited for clinical protocols because recent advances in nucleic acid chemistry allow higher cell uptake with very low off-target effects leading to actual AS-ODNs clinical applications. By using combined treatments with multiple targets it will be possible to apply TNAs directly to the cancerous cervix to destroy viral RNA and obliterate the tumor.
Single-cell RNA sequencing (scRNAseq) technology is still relatively new in the field of gastric cancer immunology but gaining significant traction. This technology now provides unprecedented ...insights into the intratumoral and intertumoral heterogeneities at the immunological, cellular, and molecular levels. Within the last few years, a volume of publications reported the usefulness of scRNAseq technology in identifying thus far elusive immunological mechanisms that may promote and impede gastric cancer development. These studies analyzed datasets generated from primary human gastric cancer tissues, metastatic ascites fluid from gastric cancer patients, and laboratory-generated data from
in vitro
and
in vivo
models of gastric diseases. In this review, we overview the exciting findings from scRNAseq datasets that uncovered the role of critical immune cells, including T cells, B cells, myeloid cells, mast cells, ILC2s, and other inflammatory stromal cells, like fibroblasts and endothelial cells. In addition, we also provide a synopsis of the initial scRNAseq findings on the interesting epithelial cell responses to inflammation. In summary, these new studies have implicated roles for T and B cells and subsets like NKT cells in tumor development and progression. The current studies identified diverse subsets of macrophages and mast cells in the tumor microenvironment, however, additional studies to determine their roles in promoting cancer growth are needed. Some groups specifically focus on the less prevalent ILC2 cell type that may contribute to early cancer development. ScRNAseq analysis also reveals that stromal cells, e.g., fibroblasts and endothelial cells, regulate inflammation and promote metastasis, making them key targets for future investigations. While evaluating the outcomes, we also highlight the gaps in the current findings and provide an assessment of what this technology holds for gastric cancer research in the coming years. With scRNAseq technology expanding rapidly, we stress the need for periodic review of the findings and assess the available scRNAseq analytical tools to guide future work on immunological mechanisms of gastric carcinogenesis.
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