DERL2 (derlin 2) stabilizes BAG6 (BAG cochaperone 6) in chemotherapy resistance of cholangiocarcinoma

DERL2 (derlin 2) is a critical component of the endoplasmic reticulum quality control pathway system whose mutations play an important role in carcinogenesis, including cholangiocarcinoma (CHOL). However, its role and its underlying mechanism have yet to be elucidated. Herein, we revealed that DERL2 was highly expressed in CHOL and considered as an independent prognostic indicator for inferior survival in CHOL. DERL2 ectopically expressed in CHOL cells promoted cell proliferation and colony formation rates, and depleting DERL2 in CHOL cells curbed tumor growth in vitro and in vivo. More interestingly, the knockout of DERL2 augmented the growth-inhibitory effect of gemcitabine chemotherapy on CHOL cells by inducing cell apoptosis. Mechanistically, we discovered that DERL2 interacted with BAG6 (BAG cochaperone 6), thereby extending its half-life and reinforcing the oncogenic role of BAG6 in CHOL progression. (AU)

DERL2 (derlin 2) stabilizes BAG6 (BAG cochaperone 6) in chemotherapy resistance of cholangiocarcinoma

DERL2 (derlin 2) is a critical component of the endoplasmic reticulum quality control pathway system whose mutations play an important role in carcinogenesis, including cholangiocarcinoma (CHOL). However, its role and its underlying mechanism have yet to be elucidated. Herein, we revealed that DERL2 was highly expressed in CHOL and considered as an independent prognostic indicator for inferior survival in CHOL. DERL2 ectopically expressed in CHOL cells promoted cell proliferation and colony formation rates, and depleting DERL2 in CHOL cells curbed tumor growth in vitro and in vivo. More interestingly, the knockout of DERL2 augmented the growth-inhibitory effect of gemcitabine chemotherapy on CHOL cells by inducing cell apoptosis. Mechanistically, we discovered that DERL2 interacted with BAG6 (BAG cochaperone 6), thereby extending its half-life and reinforcing the oncogenic role of BAG6 in CHOL progression. (AU)

DERL2 (derlin 2) stabilizes BAG6 (BAG cochaperone 6) in chemotherapy resistance of cholangiocarcinoma.

DERL2 (derlin 2) is a critical component of the endoplasmic reticulum quality control pathway system whose mutations play an important role in carcinogenesis, including cholangiocarcinoma (CHOL). However, its role and its underlying mechanism have yet to be elucidated. Herein, we revealed that DERL2 was highly expressed in CHOL and considered as an independent prognostic indicator for inferior survival in CHOL. DERL2 ectopically expressed in CHOL cells promoted cell proliferation and colony formation rates, and depleting DERL2 in CHOL cells curbed tumor growth in vitro and in vivo. More interestingly, the knockout of DERL2 augmented the growth-inhibitory effect of gemcitabine chemotherapy on CHOL cells by inducing cell apoptosis. Mechanistically, we discovered that DERL2 interacted with BAG6 (BAG cochaperone 6), thereby extending its half-life and reinforcing the oncogenic role of BAG6 in CHOL progression.

Sniffing Like a Wine Taster: Multiple Overlapping Sniffs (MOSS) Strategy Enhances Electronic Nose Odor Recognition Capability.

As highly promising devices for odor recognition, current electronic noses are still not comparable to human olfaction due to the significant disparity in the number of gas sensors versus human olfactory receptors. Inspired by the sniffing skills of wine tasters to achieve better odor perception, a multiple overlapping sniffs (MOSS) strategy is proposed in this study. The MOSS strategy involves rapid and continuous inhalation of odorants to stimulate the sensor array to generate feature-rich temporal signals. Computational fluid dynamics simulations are performed to reveal the mechanism of complex dynamic flows affecting transient responses. The proposed strategy shows over 95% accuracy in the recognition experiments of three gaseous alkanes and six liquors. Results demonstrate that the MOSS strategy can accurately and easily recognize odors with a limited sensor number. The proposed strategy has potential applications in various odor recognition scenarios, such as medical diagnosis, food quality assessment, and environmental surveillance.

Epicutaneous immunization with modified vaccinia Ankara viral vectors generates superior T cell immunity against a respiratory viral challenge.

Modified Vaccinia Ankara (MVA) was recently approved as a smallpox vaccine. Variola is transmitted by respiratory droplets and MVA immunization by skin scarification (s.s.) protected mice far more effectively against lethal respiratory challenge with vaccinia virus (VACV) than any other route of delivery, and at lower doses. Comparisons of s.s. with intradermal, subcutaneous, or intramuscular routes showed that MVAOVA s.s.-generated T cells were both more abundant and transcriptionally unique. MVAOVA s.s. produced greater numbers of lung Ova-specific CD8+ TRM and was superior in protecting mice against lethal VACVOVA respiratory challenge. Nearly as many lung TRM were generated with MVAOVA s.s. immunization compared to intra-tracheal immunization with MVAOVA and both routes vaccination protected mice against lethal pulmonary challenge with VACVOVA. Strikingly, MVAOVA s.s.-generated effector T cells exhibited overlapping gene transcriptional profiles to those generated via intra-tracheal immunization. Overall, our data suggest that heterologous MVA vectors immunized via s.s. are uniquely well-suited as vaccine vectors for respiratory pathogens, which may be relevant to COVID-19. In addition, MVA delivered via s.s. could represent a more effective dose-sparing smallpox vaccine.

Identification of an extracellular vesicle-related gene signature in the prediction of pancreatic cancer clinical prognosis.

Although extracellular vesicles (EVs) in body fluid have been considered to be ideal biomarkers for cancer diagnosis and prognosis, it is still difficult to distinguish EVs derived from tumor tissue and normal tissue. Therefore, the prognostic value of tumor-specific EVs was evaluated through related molecules in pancreatic tumor tissue. NA sequencing data of pancreatic adenocarcinoma (PAAD) were acquired from The Cancer Genome Atlas (TCGA) and International Cancer Genome Consortium (ICGC). EV-related genes in pancreatic cancer were obtained from exoRBase. Protein-protein interaction (PPI) network analysis was used to identify modules related to clinical stage. CIBERSORT was used to assess the abundance of immune and non-immune cells in the tumor microenvironment. A total of 12 PPI modules were identified, and the 3-PPI-MOD was identified based on the randomForest package. The genes of this model are involved in DNA damage and repair and cell membrane-related pathways. The independent external verification cohorts showed that the 3-PPI-MOD can significantly classify patient prognosis. Moreover, compared with the model constructed by pure gene expression, the 3-PPI-MOD showed better prognostic value. The expression of genes in the 3-PPI-MOD had a significant positive correlation with immune cells. Genes related to the hypoxia pathway were significantly enriched in the high-risk tumors predicted by the 3-PPI-MOD. External databases were used to verify the gene expression in the 3-PPI-MOD. The 3-PPI-MOD had satisfactory predictive performance and could be used as a prognostic predictive biomarker for pancreatic cancer.

Development and Validation of a Novel 8 Immune Gene Prognostic Signature Based on the Immune Expression Profile for Hepatocellular Carcinoma.

BACKGROUND: The immune microenvironment plays a vital role in the development of hepatocellular carcinoma (HCC). This study explored novel immune-related biomarkers to predict the prognosis of patients with HCC. METHODS: RNA-Seq data were downloaded from The Cancer Genome Atlas (TCGA). Univariate Cox regression was used to identify prognosis-related genes; the Lasso method was used to construct the prognosis risk model. Validation was performed on the International Cancer Genome Consortium (ICGC) cohort, and the C-index was calculated to evaluate its overall predictive performance. Western blots were conducted to evaluate the expression of genes. RESULTS: There were 320 immune-related genes, 40 of which were significantly related to prognosis. Eight immune gene signatures (CKLF, IL12A, CCL20, PRELID1, GLMN, ACVR2A, CD7, and FYN) were established by Lasso Cox regression analysis. This immune signature performed well in different cohorts and can be an independent risk factor for prognosis. In addition, the overall predictive performance of this model was higher than the other models reported previously. CONCLUSION: The predictive immune model will enable patients with HCC to be more accurately managed in immunotherapy.

In Situ H2O Meter by Visualization in Hydrogels.

The solvent content strongly affects the viscoelastic properties and network structure of hydrogels. Because of the gels' structural susceptibility and autofluorescence background, there is still no visual method to evaluate the water content in micropores. Herein, a colorimetric molecular probe (DHBYD) was synthesized for in situ visualization of water content in the micropores of hydrogels. The rapid and reversible colorimetric responses of DHBYD to solvents were obtained, which resulted a full linearity range (0 to 100%) for detecting water content in real time. Demonstrated by theoretical calculations, the sensing was attributed to changes in intramolecular charge transfer via deprotonation of phenol group. A cubic polynomial, on correlation of RGB values with water content, was established for real detection of water content in hydrogels. It reveals a new pathway for simple, in situ, and full-range evaluation of solvent content in micropores of hydrogels without any complicated procedures or expensive instruments. This would achieve fast and in situ monitoring of hydrogels to improve gel properties for better applications. It can be extended to evaluate the solvent content in other fields such as synthesis and industrial applications.

T-cell trafficking plays an essential role in tumor immunity.

Distinct populations of effector memory T cells use different homing receptors to traffic to the skin and gut. Whether tissue-selective T cells are needed for early rejection of a neoplasm growing in these tissues remains an open question. We chose to study an allogeneic tumor model because growth of such a fully mismatched tumor would signify a profound immune deficit. We implanted allogeneic tumor cells in the skin or gut of mice deficient in either α(1,3) fucosyltransferases IV and VII, enzymes critical for generating E-selectin ligands on skin-homing T cells, or ß7 integrin, a component of the α4ß7 integrin ligand for the mucosal adressin MAdCAM. During the first 9 days after tumor implantation, FucTVII-/- mice showed a profoundly impaired capacity to reject tumors growing in the skin, but readily rejected tumors implanted in the gut. Rejection of tumors in the skin was even more impaired in mice deficient in both FucTIV and FucTVII. This impairment was corrected by infusion of T cells from normal mice. By contrast, ß7 integrin-/- mice showed profoundly impaired rejection of tumors in the gut, but no defect in the skin tumor rejection. These differences were unrelated to antigen recognition or effector function of T cells, since all strains of mice were capable of generating tumor-specific CTLs in vitro against the tumor cell line used in vivo. These results demonstrate that T-cell homing defects in vivo impair immune surveillance of peripheral epithelial tissues in a specific and selective fashion.

Eccrine Syringofibroadenoma in Association With Acquired Epidermodysplasia Verruciformis.

A 75-year-old man with human immunodeficiency virus infection and numerous biopsy-proven warts for 10 years, refractory to cryosurgery, cimetidine, and topical imiquimod, presented with numerous pink to hypopigmented verrucous papules and plaques involving the face, trunk, buttocks, and groin. Laboratory evaluation revealed a CD4 T-cell count of 62 cells per microliter and human immunodeficiency virus viral load of <117 copies per milliliter. Biopsy of a plaque groin lesion was performed. Histopathology revealed vertically oriented anastomosing strands of basaloid epithelium arising from multiple points along the epidermis in a background fibrovascular stroma. Ductal differentiation was identified. Areas of epidermis showed compact orthokeratosis, coarse hypergranulosis, and keratinocytes with abundant steel-blue-gray cytoplasm, indicative of viral cytopathic changes. Cytologic atypia was not identified. Human papillomavirus (HPV) genotyping of this lesion was positive for types 5 and 14. Overall, the findings were consistent with epidermodysplasia verruciformis in association with eccrine syringofibroadenoma (ESFA). The patient was subsequently treated with acitretin and showed clinical improvement. ESFA is an uncommon benign adnexal tumor with unknown pathogenesis. Although its association with HPV has rarely been reported, ESFA in the setting of acquired epidermodysplasia verruciformis has not been described. The development of ESFA in this case may be the result of HPV-induced cellular transformation.

Skin infection generates non-migratory memory CD8+ T(RM) cells providing global skin immunity.

Protective T-cell memory has long been thought to reside in blood and lymph nodes, but recently the concept of immune memory in peripheral tissues mediated by resident memory T (T(RM)) cells has been proposed. Here we show in mice that localized vaccinia virus (VACV) skin infection generates long-lived non-recirculating CD8(+) skin T(RM) cells that reside within the entire skin. These skin T(RM) cells are potent effector cells, and are superior to circulating central memory T (T(CM)) cells at providing rapid long-term protection against cutaneous re-infection. We find that CD8(+) T cells are rapidly recruited to skin after acute VACV infection. CD8(+) T-cell recruitment to skin is independent of CD4(+) T cells and interferon-γ, but requires the expression of E- and P-selectin ligands by CD8(+) T cells. Using parabiotic mice, we further show that circulating CD8(+) T(CM) and CD8(+) skin T(RM) cells are both generated after skin infection; however, CD8(+) T(CM) cells recirculate between blood and lymph nodes whereas T(RM) cells remain in the skin. Cutaneous CD8(+) T(RM) cells produce effector cytokines and persist for at least 6 months after infection. Mice with CD8(+) skin T(RM) cells rapidly cleared a subsequent re-infection with VACV whereas mice with circulating T(CM) but no skin T(RM) cells showed greatly impaired viral clearance, indicating that T(RM) cells provide superior protection. Finally, we show that T(RM) cells generated as a result of localized VACV skin infection reside not only in the site of infection, but also populate the entire skin surface and remain present for many months. Repeated re-infections lead to progressive accumulation of highly protective T(RM) cells in non-involved skin. These findings have important implications for our understanding of protective immune memory at epithelial interfaces with the environment, and suggest novel strategies for vaccines that protect against tissue tropic organisms.

Disruption of TNF-α/TNFR1 function in resident skin cells impairs host immune response against cutaneous vaccinia virus infection.

One strategy adopted by vaccinia virus (VV) to evade the host immune system is to encode homologs of TNF receptors (TNFRs) that block TNF-α function. The response to VV skin infection under conditions of TNF-α deficiency, however, has not been reported. We found that TNFR1-/- mice developed larger primary lesions, numerous satellite lesions, and higher skin virus levels after VV scarification. Following their recovery, VV-scarified TNFR1-/- mice were fully protected against challenge with a lethal intranasal dose of VV, suggesting these mice had developed an effective memory immune response. A functional systemic immune response was further demonstrated by enhanced production of VV-specific IFN-γ and VV-specific CD8(+) T cells in spleens and draining lymph nodes. Interestingly, bone marrow (BM)-reconstitution studies using wild-type (WT) BM in TNFR1-/- host mice, but not TNFR1-/- BM in WT host mice, reproduced the original results seen in TNFR1-/- mice, indicating that TNFR1 deficiency in resident skin cells, rather than hematopoietic cells, accounts for the impaired cutaneous immune response. Our data suggest that lack of TNFR1 leads to a skin-specific immune deficiency, and that resident skin cells have a crucial role in mediating an optimal immune defense to VV cutaneous infection via TNF-α/TNFR1 signaling.

Epidermal injury and infection during poxvirus immunization is crucial for the generation of highly protective T cell-mediated immunity.

Variola major (smallpox) infection claimed hundreds of millions lives before it was eradicated by a simple vaccination strategy: epicutaneous application of the related orthopoxvirus vaccinia virus (VACV) to superficially injured skin (skin scarification, s.s.). However, the remarkable success of this strategy was attributed to the immunogenicity of VACV rather than to the unique mode of vaccine delivery. We now show that VACV immunization via s.s., but not conventional injection routes, is essential for the generation of superior T cell-mediated immune responses that provide complete protection against subsequent challenges, independent of neutralizing antibodies. Skin-resident effector memory T cells (T(EM) cells) provide complete protection against cutaneous challenge, whereas protection against lethal respiratory challenge requires both respiratory mucosal T(EM) cells and central memory T cells (T(CM) cells). Vaccination with recombinant VACV (rVACV) expressing a tumor antigen was protective against tumor challenge only if delivered via the s.s. route; it was ineffective if delivered by hypodermic injection. The clinically safer nonreplicative modified vaccinia Ankara virus (MVA) also generated far superior protective immunity when delivered via the s.s. route compared to intramuscular (i.m.) injection as used in MVA clinical trials. Thus, delivery of rVACV-based vaccines, including MVA vaccines, through physically disrupted epidermis has clear-cut advantages over conventional vaccination via hypodermic injection.

Overexpression of IL-1alpha in skin differentially modulates the immune response to scarification with vaccinia virus.

Transepidermal inoculation of vaccinia virus (VV), or scarification, has been used effectively for the induction of specific and long-lasting immunity to smallpox and is superior to other routes of immunization. Scarification of individuals with atopic skin disease or immune deficiency, however, can lead to persistent viral replication and result in significant morbidity and mortality. These effects of scarification presumably reflect the unique immunological properties of skin and the immune cells resident in, or recruited to, the site of inoculation. To explore these phenomena, we utilized transgenic mice engineered to overexpress IL-1alpha, a critical mediator of cutaneous inflammation, in the epidermis. Following scarification with VV, both transgenic and wild-type mice develop local pox. At high doses of VV, IL-1alpha transgenic mice recruited immune cells to the inoculation site more rapidly and demonstrated enhanced T-cell and humoral immune responses. At limiting doses, however, IL-1alpha transgenic mice could effectively control virus replication without formation of pox lesions or activation of a memory response. This study suggests that IL-1 might be useful as an adjuvant to enhance antiviral immunity and promote safer vaccination strategies; however, understanding the balance of IL-1 effects on innate and adaptive immune functions will be critical to achieve optimal results.

Dendritic cells are preferentially targeted among hematolymphocytes by Modified Vaccinia Virus Ankara and play a key role in the induction of virus-specific T cell responses in vivo.

BACKGROUND: Modified Vaccinia Ankara (MVA) is a highly attenuated strain of vaccinia virus (VV) that has lost approximately 15% of the VV genome, along with the ability to replicate in most mammalian cells. It has demonstrated impressive safety and immunogenicity profile in both preclinical and clinical studies, and is being actively explored as a promising vaccine vector for a number of infectious diseases and malignancies. However, little is known about how MVA interacts with the host immune system constituents, especially dendritic cells (DCs), to induce strong immune responses despite its inability to replicate in vivo. Using in vitro and in vivo murine models, we systematically investigated the susceptibility of murine DCs to MVA infection, and the immunological consequences of the infection. RESULTS: Our data demonstrate that MVA preferentially infects professional antigen presenting cells, especially DCs, among all the subsets of hematolymphoid cells. In contrast to the reported blockage of DC maturation and function upon VV infection, DCs infected by MVA undergo phenotypic maturation and produce innate cytokine IFN-alpha within 18 h of infection. Substantial apoptosis of MVA-infected DCs occurs after 12 h following infection and the apoptotic DCs are readily phagocytosed by uninfected DCs. Using MHC class I - deficient mice, we showed that both direct and cross-presentation of viral Ags are likely to be involved in generating viral-specific CD8+ T cell responses. Finally, DC depletion abrogated the T cell activation in vivo. CONCLUSION: We present the first in vivo evidence that among hematolymphoid cells, DCs are the most susceptible targets for MVA infection, and DC-mediated Ag presentation is required for the induction of MVA-specific immune responses. These results provide important information concerning the mechanisms by which strong immune responses are elicited to MVA-encoded antigens and may inform efforts to further improve the immunogenicity of this already promising vaccine vector.

Skin-homing receptors on effector leukocytes are differentially sensitive to glyco-metabolic antagonism in allergic contact dermatitis.

T cell recruitment into inflamed skin is dependent on skin-homing receptor binding to endothelial (E)- and platelet (P)-selectin. These T cell receptors, or E- and P-selectin ligands, can be targeted by the metabolic fluorosugar inhibitor, 4-F-GlcNAc, to blunt cutaneous inflammation. Compelling new data indicate that, in addition to T cells, NK cells are also recruited to inflamed skin in allergic contact hypersensitivity (CHS) contingent on E- and P-selectin-binding. Using a model of allergic CHS, we evaluated the identity and impact of NK cell E-selectin ligand(s) on inflammatory responses and examined the oral efficacy of 4-F-GlcNAc. We demonstrated that the predominant E-selectin ligands on NK cells are P-selectin glycoprotein ligand-1 and protease-resistant glycolipids. We showed that, unlike the induced E-selectin ligand expression on activated T cells upon exposure to Ag, ligand expression on NK cells was constitutive. CHS responses were significantly lowered by orally administered 4-F-GlcNAc treatment. Although E-selectin ligand on activated T cells was suppressed, ligand expression on NK cells was insensitive to 4-F-GlcNAc treatment. These findings indicate that downregulating effector T cell E- and P-selectin ligand expression directly correlates with anti-inflammatory efficacy and provides new insight on metabolic discrepancies of E-selectin ligand biosynthesis in effector leukocytes in vivo.

Dynamic programming of CD8+ T cell trafficking after live viral immunization.

After viral infection, activated T cells are present in multiple tissues regardless of the infection route. How these cells acquire pleiotropic homing ability is unclear. By using a cutaneous vaccinia virus infection model, we demonstrate that regulation of T cell trafficking is multiphasic. Upon completion of three cell divisions, CD8+ T cells upregulated specific skin-homing molecules within draining lymph nodes (LN). By 60 hr after infection, some activated T cells reached the infected tissue, while others entered distant antigen-free LN. These latter cells continued to divide and acquire additional tissue-homing molecules in this new setting, independent of antigen presentation. After viral clearance, the initial skin-homing imprint became the predominant homing phenotype on memory cells and provided superior protection against secondary cutaneous challenge. These observations demonstrate a mechanism by which T cells provide both immediate tissue-specific immune control at the pathogen entry site and a more flexible systemic protection against pathogen dissemination.

The majority of human peripheral blood CD4+CD25highFoxp3+ regulatory T cells bear functional skin-homing receptors.

CD4+CD25+ T regulatory cells (Treg) are thought to be important in the peripheral tolerance. Recent evidence suggests that human peripheral blood CD4+CD25+ T cells are heterogeneous and contain both CD4+CD25(high) T cells with potent regulatory activity and many more CD4+CD25(low/med) nonregulatory T cells. In this study, we found that virtually all peripheral blood CD4+CD25(high)Foxp3+ Treg expressed high levels of the chemokine receptor CCR4. In addition, 80% of Treg expressed cutaneous lymphocyte Ag (CLA) and 73% expressed CCR6. These molecules were functional, as CLA+ Treg showed CD62E ligand activity and demonstrable chemotactic responses to the CCR4 ligands CCL22 and CCL17 and to the CCR6 ligand CCL20. The phenotype and chemotactic response of these Treg were significantly different from those of CD4+CD25(med) nonregulatory T cells. We further demonstrated that blood CLA+ Treg inhibited CD4+CD25- T cell proliferation induced by anti-CD3. Based on homing receptor profile, CLA+ Treg should enter normal skin. We next isolated CD4+CD25(high) T cells directly from normal human skin; these cells suppressed proliferation of skin CD4+CD25- T cells. Therefore, the majority of true circulating Treg express functional skin-homing receptors, and human Treg may regulate local immune responses in normal human skin.