Development and validation of a major adverse limb events prediction model for peripheral arterial disease with frailty.

OBJECTIVE: To investigate the risk factors for major limb adverse events (MALE) in peripheral arterial disease (PAD) combined with frailty and to develop and validate a risk prediction model of MALE. METHODS: This prospective study was performed in the vascular surgery department of patients in six hospitals in southwest China. Prospective collection of patients with PAD combined with frailty from February 1 to December 20, 2021, with MALE as the primary outcome, and followed for 1 year. The cohort was divided into a development cohort and a validation cohort. In the development cohort, a multivariate risk prediction model was developed to predict MALE using random forests for variable selection and multivariable Cox regression analysis. The model is represented by a visualized nomogram and a web-based calculator. The model performance was tested with the validation cohort and assessed using the C-statistic and calibration plots. RESULTS: A total of 1179 patients were prospectively enrolled from February 1 to December 20, 2021. Among 816 patients with PAD who were included in the analysis, the median follow-up period for this study was 9 ± 4.07 months, the mean age was 74.64 ± 9.43 years, and 249 (30.5%) were women. Within 1 year, 222 patients (27.2%) developed MALE. Target lesion revascularizations were performed in 99 patients (12.1%), and amputations were performed in 131 patients (16.1%). The mortality rate within the whole cohort was 108 patients (13.2%). After controlling for competing risk events (death), the cumulative risk of developing MALE was not statistically different. Prealbumin (hazard ratio [HR], 0.6; 95% confidence interval [CI], 0.41-0.89; P = .010), percutaneous coronary intervention (HR, 2.31; 95% CI, 1.26-4.21; P = .006), Rutherford classification (HR, 1.77; 95% CI, 1.36-2.31; P < .001), white blood cell (HR, 1.85; 95% CI, 1.20-2.87; P = .005), high altitude area (HR, 3.1; 95% CI, 1.43-6.75; P = .004), endovascular treatment (HR, 10.2; 95% CI, 1.44-72.50; P = .020), and length of stay (HR, 1.01; 95% CI, 1.00-1.03; P = .012) were risk factors for MALE. The MALE prediction model had a C-statistic of 0.76 (95% CI, 0.70-0.79). The C-statistic was 0.68 for internal validation and 0.66 for external validation for the MALE prediction model. The MALE prediction model for PAD presented an interactive nomogram and a web-based network calculator. CONCLUSIONS: In this study, the MALE prediction model has a discriminative ability to predict MALE among patients with PAD in frailty. The MALE model can optimize clinical decision-making for patients with PAD in frailty.

Transcriptional regulation by NR5A2 links differentiation and inflammation in the pancreas.

Chronic inflammation increases the risk of developing one of several types of cancer. Inflammatory responses are currently thought to be controlled by mechanisms that rely on transcriptional networks that are distinct from those involved in cell differentiation. The orphan nuclear receptor NR5A2 participates in a wide variety of processes, including cholesterol and glucose metabolism in the liver, resolution of endoplasmic reticulum stress, intestinal glucocorticoid production, pancreatic development and acinar differentiation. In genome-wide association studies, single nucleotide polymorphisms in the vicinity of NR5A2 have previously been associated with the risk of pancreatic adenocarcinoma. In mice, Nr5a2 heterozygosity sensitizes the pancreas to damage, impairs regeneration and cooperates with mutant Kras in tumour progression. Here, using a global transcriptomic analysis, we describe an epithelial-cell-autonomous basal pre-inflammatory state in the pancreas of Nr5a2+/- mice that is reminiscent of the early stages of pancreatitis-induced inflammation and is conserved in histologically normal human pancreases with reduced expression of NR5A2 mRNA. In Nr5a2+/-mice, NR5A2 undergoes a marked transcriptional switch, relocating from differentiation-specific to inflammatory genes and thereby promoting gene transcription that is dependent on the AP-1 transcription factor. Pancreatic deletion of Jun rescues the pre-inflammatory phenotype, as well as binding of NR5A2 to inflammatory gene promoters and the defective regenerative response to damage. These findings support the notion that, in the pancreas, the transcriptional networks involved in differentiation-specific functions also suppress inflammatory programmes. Under conditions of genetic or environmental constraint, these networks can be subverted to foster inflammation.

Facile Synthesis of ZrO2-SiO2 Antireflective Films with Good Mechanical Performances for Perovskite Solar Cells.

Antireflective (AR) films are widely applied in solar cells to reduce the reflectivity toward sunlight, thus improving the photoelectric conversion efficiency (PCE) of solar cells. However, AR films are still suffering from poor mechanical properties and low transmittance in photovoltaic applications. Herein, a ZrO2-SiO2 composite film with enhanced mechanical properties was successfully synthesized by a facile sol-gel method, whose pencil hardness increased from less than 6B to B compared with the pure SiO2 film synthesized with the same alkali-catalyzed method. Moreover, the ZrO2-SiO2 film with a Zr/Si mole ratio (nZr/Si) of 0.06 exhibited a high transmittance gain (ΔT) of 3.0%, and an obvious increase (1.32%) in PCE was observed in a perovskite solar cell compared with the cell covered by a bare glass. Additionally, both the short-circuit current density (JSC) and PCE of perovskite solar cells have a non-linear increasing relationship with the average transmittance (Tavg) of the ZrO2-SiO2 composite film. In this sense, this work can provide a facile way to prepare AR films effectively improving performances of solar cells.

Reversal of autoimmunity by mixed chimerism enables reactivation of ß cells and transdifferentiation of α cells in diabetic NOD mice.

Type 1 diabetes (T1D) results from the autoimmune destruction of ß cells, so cure of firmly established T1D requires both reversal of autoimmunity and restoration of ß cells. It is known that ß cell regeneration in nonautoimmune diabetic mice can come from differentiation of progenitors and/or transdifferentiation of α cells. However, the source of ß cell regeneration in autoimmune nonobese diabetic (NOD) mice remains unclear. Here, we show that, after reversal of autoimmunity by induction of haploidentical mixed chimerism, administration of gastrin plus epidermal growth factor augments ß cell regeneration and normalizes blood glucose in the firmly established diabetic NOD mice. Using transgenic NOD mice with inducible lineage-tracing markers for insulin-producing ß cells, Sox9+ ductal progenitors, Nestin+ mesenchymal stem cells, and glucagon-producing α cells, we have found that both reactivation of dysfunctional low-level insulin expression (insulinlo) ß cells and neogenesis contribute to the regeneration, with the latter predominantly coming from transdifferentiation of α cells. These results indicate that, after reversal of autoimmunity, reactivation of ß cells and transdifferentiation of α cells can provide sufficient new functional ß cells to reach euglycemia in firmly established T1D.

Targeting disordered-structured domain interactions in Galectin-3 based on NMR and enhanced MD.

Intrinsically disordered regions (IDRs) are common and important functional domains in many proteins. However, IDRs are difficult to target for drug development due to the lack of defined structures that would facilitate the identification of possible drug-binding pockets. Galectin-3 is a carbohydrate-binding protein of which overexpression has been implicated in a wide variety of disorders, including cancer and inflammation. Apart from its carbohydrate-recognition/binding domain (CRD), Galectin-3 also contains a functionally important disordered N-terminal domain (NTD) that contacts the C-terminal domain (CTD) and could be a target for drug development. To overcome challenges involved in inhibitor design due to lack of structure and the highly dynamic nature of the NTD, we used a protocol combining nuclear magnetic resonance data from recombinant Galectin-3 with accelerated molecular dynamics (MD) simulations. This approach identified a pocket in the CTD with which the NTD makes frequent contact. In accordance with this model, mutation of residues L131 and L203 in this pocket caused loss of Galectin-3 agglutination ability, signifying the functional relevance of the cavity. In silico screening was used to design candidate inhibitory peptides targeting the newly discovered cavity, and experimental testing of only three of these yielded one peptide that inhibits the agglutination promoted by wild-type Galectin-3. NMR experiments further confirmed that this peptide indeed binds to a cavity in the CTD, not within the actual CRD. Our results show that it is possible to apply a combination of MD simulations and NMR experiments to precisely predict the binding interface of a disordered domain with a structured domain, and furthermore use this predicted interface for designing inhibitors. This procedure can potentially be extended to many other targets in which similar IDR interactions play a vital functional role.

Observing Mesoscopic Nucleic Acid Capacitance Effect and Mismatch Impact via Graphene Transistors.

This work reports a molecular-scale capacitance effect of the double helical nucleic acid duplex structure for the first time. By quantitatively conducting large sample measurements of the electrostatic field effect using a type of high-accuracy graphene transistor biosensor, an unusual charge-transport behavior is observed in which the end-immobilized nucleic acid duplexes can store a part of ionization electrons like molecular capacitors, other than electric conductors. To elucidate this discovery, a cascaded capacitive network model is proposed as a novel equivalent circuit of nucleic acid duplexes, expanding the point-charge approximation model, by which the partial charge-transport observation is reasonably attributed to an electron-redistribution behavior within the capacitive network. Furthermore, it is experimentally confirmed that base-pair mismatches hinder the charge transport in double helical duplexes, and lead to directly identifiable alterations in electrostatic field effects. The bioelectronic principle of mismatch impact is also self-consistently explained by the newly proposed capacitive network model. The mesoscopic nucleic acid capacitance effect may enable a new kind of label-free nucleic acid analysis tool based on electronic transistor devices. The in situ and real-time nucleic acid detections for virus biomarkers, somatic mutations, and genome editing off-target may thus be predictable.

Galectin-1 and Galectin-3 in B-Cell Precursor Acute Lymphoblastic Leukemia.

Acute lymphoblastic leukemias arising from the malignant transformation of B-cell precursors (BCP-ALLs) are protected against chemotherapy by both intrinsic factors as well as by interactions with bone marrow stromal cells. Galectin-1 and Galectin-3 are lectins with overlapping specificity for binding polyLacNAc glycans. Both are expressed by bone marrow stromal cells and by hematopoietic cells but show different patterns of expression, with Galectin-3 dynamically regulated by extrinsic factors such as chemotherapy. In a comparison of Galectin-1 x Galectin-3 double null mutant to wild-type murine BCP-ALL cells, we found reduced migration, inhibition of proliferation, and increased sensitivity to drug treatment in the double knockout cells. Plant-derived carbohydrates GM-CT-01 and GR-MD-02 were used to inhibit extracellular Galectin-1/-3 binding to BCP-ALL cells in co-culture with stromal cells. Treatment with these compounds attenuated migration of the BCP-ALL cells to stromal cells and sensitized human BCP-ALL cells to vincristine and the targeted tyrosine kinase inhibitor nilotinib. Because N-glycan sialylation catalyzed by the enzyme ST6Gal1 can regulate Galectin cell-surface binding, we also compared the ability of BCP-ALL wild-type and ST6Gal1 knockdown cells to resist vincristine treatment when they were co-cultured with Galectin-1 or Galectin-3 knockout stromal cells. Consistent with previous results, stromal Galectin-3 was important for maintaining BCP-ALL fitness during chemotherapy exposure. In contrast, stromal Galectin-1 did not significantly contribute to drug resistance, and there was no clear effect of ST6Gal1-catalysed N-glycan sialylation. Taken together, our results indicate a complicated joint contribution of Galectin-1 and Galectin-3 to BCP-ALL survival, with different roles for endogenous and stromal produced Galectins. These data indicate it will be important to efficiently block both extracellular and intracellular Galectin-1 and Galectin-3 with the goal of reducing BCP-ALL persistence in the protective bone marrow niche during chemotherapy.

SINR- and MI-Based Double-Robust Waveform Design.

Owing to cognitive radar breaking the open-loop receiving-transmitting mode of traditional radar, adaptive waveform design for cognitive radar has become a central issue in radar system research. In this paper, the method of radar transmitted waveform design in the presence of clutter is studied. Since exact characterizations of the target and clutter spectra are uncommon in practice, a single-robust transmitted waveform design method is introduced to solve the problem of the imprecise target spectrum or the imprecise clutter spectrum. Furthermore, considering that radar cannot simultaneously obtain precise target and clutter spectra, a novel double-robust transmitted waveform design method is proposed. In this method, the signal-to-interference-plus-noise ratio and mutual information are used as the objective functions, and the optimization models for the double-robust waveform are established under the transmitted energy constraint. The Lagrange multiplier method was used to solve the optimal double-robust transmitted waveform. The simulation results show that the double-robust transmitted waveform can maximize SINR and MI in the worst case; the performance of SINR and MI will degrade if other transmitted waveforms are employed in the radar system.

PGC-1 coactivators regulate MITF and the tanning response.

The production of pigment by melanocytes tans the skin and protects against skin cancers. UV-exposed keratinocytes secrete α-MSH, which then activates melanin formation in melanocytes by inducing the microphthalmia-associated transcription factor (MITF). We show that PPAR-γ coactivator (PGC)-1α and PGC-1ß are critical components of this melanogenic system in melanocytes. α-MSH signaling strongly induces PGC-1α expression and stabilizes both PGC-1α and PGC-1ß proteins. The PGC-1s in turn activate the MITF promoter, and their expression correlates strongly with that of MITF in human melanoma cell lines and biopsy specimens. Inhibition of PGC-1α and PGC-1ß blocks the α-MSH-mediated induction of MITF and melanogenic genes. Conversely, overexpression of PGC-1α induces pigment formation in cell culture and transgenic animals. Finally, polymorphism studies reveal expression quantitative trait loci in the PGC-1ß gene that correlate with tanning ability and protection from melanoma in humans. These data identify PGC-1 coactivators as regulators of human tanning.

Are preventive measures adequate? An evaluation of the implementation of COVID-19 prevention and control measures in nursing homes in China.

BACKGROUND: The novel coronavirus disease 2019 (COVID-19) pandemic has become a challenge for nursing homes in China. Nursing homes are particularly dangerous places in terms of the spread of COVID-19 given that they house vulnerable, high-risk populations. As such, several useful guidelines for coping with COVID-19 in nursing homes have been provided. However, the actual implementation rates of such guidelines are unknown. This study aims to document the adherence of nursing homes to the Ministry of Civil Affairs guidelines for COVID-19 prevention and control in nursing homes. METHODS: A cross-sectional study was conducted among 484 nursing homes in 136 cities of 28 provinces in China. A self-report questionnaire was created based on the Ministry of Civil Affairs guidelines for COVID-19 prevention and control in nursing homes (first edition). The questionnaire and the Transformational Leadership in the Public Sector Scale were sent to nursing home managers via the Wenjuanxing app online from February 7 to 29, 2020. Ultimately, 461 of 960 nursing homes participated, for a response rate of 48.0%. RESULTS: The average overall implementation rate of COVID-19 prevention and control measures was 80.0% (143.97/180). The average implementation rates for hygienic behaviour management and access management were lower, at 75.3 and 78.7%, respectively. Number of medical staff and transformational leadership score of nursing home's manager were associated with total implementation score (p < 0.05). A total of 69.8% (322/461) of the nursing home managers had serious resource problems, and inadequate protective supplies (72.0%) and staff shortages (47.7%) were the two primary problems. The nursing homes that located in urban, with large nursing home size, had hospital-nursing home cooperation and the transformational leadership score of manager> 60, had a lower risk of having serious resource problems. CONCLUSIONS: Overall, the implementation of prevention and control measures by nursing homes are insufficient during the epidemic in China. More medical staff, adequate resource, cooperation with hospitals, and higher transformational leadership of manager are required to improve the implementation rate. It is urgent for nursing homes to maintain the safety of residents and staff.

MHC-mismatched mixed chimerism restores peripheral tolerance of noncross-reactive autoreactive T cells in NOD mice.

Autoimmune type 1 diabetes (T1D) and other autoimmune diseases are associated with particular MHC haplotypes and expansion of autoreactive T cells. Induction of MHC-mismatched but not -matched mixed chimerism by hematopoietic cell transplantation effectively reverses autoimmunity in diabetic nonobese diabetic (NOD) mice, even those with established diabetes. As expected, MHC-mismatched mixed chimerism mediates deletion in the thymus of host-type autoreactive T cells that have T-cell receptor (TCR) recognizing (cross-reacting with) donor-type antigen presenting cells (APCs), which have come to reside in the thymus. However, how MHC-mismatched mixed chimerism tolerizes host autoreactive T cells that recognize only self-MHC-peptide complexes remains unknown. Here, using NOD.Rag1-/-BDC2.5 or NOD.Rag1-/-BDC12-4.1 mice that have only noncross-reactive transgenic autoreactive T cells, we show that induction of MHC-mismatched but not -matched mixed chimerism restores immune tolerance of peripheral noncross-reactive autoreactive T cells. MHC-mismatched mixed chimerism results in increased percentages of both donor- and host-type Foxp3+ Treg cells and up-regulated expression of programmed death-ligand 1 (PD-L1) by host-type plasmacytoid dendritic cells (pDCs). Furthermore, adoptive transfer experiments showed that engraftment of donor-type dendritic cells (DCs) and expansion of donor-type Treg cells are required for tolerizing the noncross-reactive autoreactive T cells in the periphery, which are in association with up-regulation of host-type DC expression of PD-L1 and increased percentage of host-type Treg cells. Thus, induction of MHC-mismatched mixed chimerism may establish a peripheral tolerogenic DC and Treg network that actively tolerizes autoreactive T cells, even those with no TCR recognition of the donor APCs.

A quantitative bias analysis of the confounding effects due to smoking on the association between fluoroquinolones and risk of aortic aneurysm.

PURPOSE: Epidemiologic studies consistently report an increased risk of aortic aneurysm (AA) among users of fluoroquinolones (FQ), but confounding by smoking could explain all or some of the observed risk. Therefore, to better elucidate the potential causal impact of FQ on AA, we quantitatively evaluated the potential confounding effect of smoking on this observed association. METHODS: We conducted a series of quantitative bias analyses using three previously published approaches: the E-value approach, the rule-out approach, and the array approach. We additionally conducted a numerical comparison between the rule-out approach and the E-value approach. RESULTS: For an apparent relative risk of 2, the E-value is 3.41, suggesting that smoking needs to be associated with both FQ and AA with a minimal magnitude of 3.41 to explain away the observed twofold FQ-AA association. The array approach found that the prevalence of smoking among FQ users would need to be at least 2.9 times higher (43%) than the nonusers (15%), assuming smoking increases the risk of AA by 7.6-fold. A numerical comparison demonstrated that the results from the rule-out approach are similar to that of the E-value approach when there is a lack of prior data on bias parameters. CONCLUSIONS: Using three different approaches, we demonstrate that the strengths of association between smoking and both FQ and AA need to be unusually strong to fully account for the twofold increased risk between FQ and AA. Therefore, it is unlikely that smoking alone would explain away the association reported in the epidemiologic studies.

MiR-3619-5p hampers proliferation and cisplatin resistance in cutaneous squamous-cell carcinoma via KPNA4.

Cutaneous squamous cell carcinoma (CSCC) remains the second most prevailing cancer worldwide and presents high mortality rates. Given that chemoresistance becomes an enormous obstacle to the therapy for CSCC patients, there is a pressing need to discover novel strategies for enhancing the response of CSCC cells to cisplatin. Emerging evidence has unfolded that miRNAs are participated in regulation of drug resistance in multiple cancers. MiR-3619-5p has been proofed to exert tumor inhibitive activities in human malignancies, but the biological function of miR-3619-5p in the progression of CSCC is still unclear. In this study, we observed that miR-3619-5p expression was pronouncedly dropped in cisplatin-resistant CSCC cells. Subsequently, miR-3619-5p was validated to act as a tumor suppressor in CSCC through retarding cell proliferation and cisplatin resistance. Besides, our findings certified that KPNA4 was highly expressed in cisplatin-resistant CSCC cells. Further, KPNA4 was negatively regulated by miR-3619-5p. Rescue experiments unveiled that KPNA4 counteracted the miR-3619-5p-mediated regulation of CSCC tumorigenesis. On the whole, miR-3619-5p inhibited cell proliferation and cisplatin resistance of CSCC by regulating KPNA4 expression, suggesting that miR-3619-5p/KPNA4 pathway may represent a potential promising strategy for the treatment of patients with CSCC.

Isoorientin alleviates UVB-induced skin injury by regulating mitochondrial ROS and cellular autophagy.

Ultraviolet B (UVB) irradiation increases the risk of various skin disorders, resulting in apoptosis, autophagy and oxidative stress and thereby promoting the risk of skin photoaging and carcinogenesis. The use of photochemoprotectors including natural products with antioxidant properties represents an effective strategy for preventing UVB-induced skin injury. Isoorientin (Iso), as a flavonoid compound, could be extracted from several plant species and possesses multiple biological activities. However, its role in regulating UVB-induced skin damage is little to be reported. In the study, we found that Iso treatment could protect human dermal fibroblasts (HDFs) against the effects of UVB irradiation by improving cell viability, suppressing MMP1 and MMP3 expression, inhibiting oxidative stress and inducing autophagy. In addition, Iso reduced UVB-triggered apoptosis, as evidenced by the decreased Caspase-3 activity in vitro. Furthermore, Iso was functioned as reactive oxygen species (ROS) scavenger that markedly hindered c-Jun N-terminal kinases (JNK) signaling activation in UVB-treated HFDs. Importantly, promoting JNK activity restored matrix metalloproteinase (MMP)-1/3 expression in Iso-incubated cells with UVB stimulation. Meanwhile, UVB exposure to the skin of mice and subsequent topical application of Iso delayed the progression of skin damage, resulting in autophagy and blocking the JNK activation and ROS production. In conclusion, these results indicated the photoprotective role of Iso and demonstrated that Iso could also be potentially used as an agent against UVB-stimulated skin damage.

Growth factors and medium hyperglycemia induce Sox9+ ductal cell differentiation into ß cells in mice with reversal of diabetes.

We previously reported that long-term administration of a low dose of gastrin and epidermal growth factor (GE) augments ß-cell neogenesis in late-stage diabetic autoimmune mice after eliminating insulitis by induction of mixed chimerism. However, the source of ß-cell neogenesis is still unknown. SRY (sex-determining region Y)-box 9(+) (Sox9(+)) ductal cells in the adult pancreas are clonogenic and can give rise to insulin-producing ß cells in an in vitro culture. Whether Sox9(+) ductal cells in the adult pancreas can give rise to ß cells in vivo remains controversial. Here, using lineage-tracing with genetic labeling of Insulin- or Sox9-expressing cells, we show that hyperglycemia (>300 mg/dL) is required for inducing Sox9(+) ductal cell differentiation into insulin-producing ß cells, and medium hyperglycemia (300-450 mg/dL) in combination with long-term administration of low-dose GE synergistically augments differentiation and is associated with normalization of blood glucose in nonautoimmune diabetic C57BL/6 mice. Short-term administration of high-dose GE cannot augment differentiation, although it can augment preexisting ß-cell replication. These results indicate that medium hyperglycemia combined with long-term administration of low-dose GE represents one way to induce Sox9(+) ductal cell differentiation into ß cells in adult mice.

Characterising cis-regulatory variation in the transcriptome of histologically normal and tumour-derived pancreatic tissues.

OBJECTIVE: To elucidate the genetic architecture of gene expression in pancreatic tissues. DESIGN: We performed expression quantitative trait locus (eQTL) analysis in histologically normal pancreatic tissue samples (n=95) using RNA sequencing and the corresponding 1000 genomes imputed germline genotypes. Data from pancreatic tumour-derived tissue samples (n=115) from The Cancer Genome Atlas were included for comparison. RESULTS: We identified 38 615 cis-eQTLs (in 484 genes) in histologically normal tissues and 39 713 cis-eQTL (in 237 genes) in tumour-derived tissues (false discovery rate <0.1), with the strongest effects seen near transcriptional start sites. Approximately 23% and 42% of genes with significant cis-eQTLs appeared to be specific for tumour-derived and normal-derived tissues, respectively. Significant enrichment of cis-eQTL variants was noted in non-coding regulatory regions, in particular for pancreatic tissues (1.53-fold to 3.12-fold, p≤0.0001), indicating tissue-specific functional relevance. A common pancreatic cancer risk locus on 9q34.2 (rs687289) was associated with ABO expression in histologically normal (p=5.8×10-8) and tumour-derived (p=8.3×10-5) tissues. The high linkage disequilibrium between this variant and the O blood group generating deletion variant in ABO (exon 6) suggested that nonsense-mediated decay (NMD) of the 'O' mRNA might explain this finding. However, knockdown of crucial NMD regulators did not influence decay of the ABO 'O' mRNA, indicating that a gene regulatory element influenced by pancreatic cancer risk alleles may underlie the eQTL. CONCLUSIONS: We have identified cis-eQTLs representing potential functional regulatory variants in the pancreas and generated a rich data set for further studies on gene expression and its regulation in pancreatic tissues.

Functional characterization of a chr13q22.1 pancreatic cancer risk locus reveals long-range interaction and allele-specific effects on DIS3 expression.

Genome-wide association studies (GWAS) have identified multiple common susceptibility loci for pancreatic cancer. Here we report fine-mapping and functional analysis of one such locus residing in a 610 kb gene desert on chr13q22.1 (marked by rs9543325). The closest candidate genes, KLF5, KLF12, PIBF1, DIS3 and BORA, range in distance from 265-586 kb. Sequencing three sub-regions containing the top ranked SNPs by imputation P-value revealed a 30 bp insertion/deletion (indel) variant that was significantly associated with pancreatic cancer risk (rs386772267, P = 2.30 × 10-11, OR = 1.22, 95% CI 1.15-1.28) and highly correlated to rs9543325 (r2 = 0.97 in the 1000 Genomes EUR population). This indel was the most significant cis-eQTL variant in a set of 222 histologically normal pancreatic tissue samples (ß = 0.26, P = 0.004), with the insertion (risk-increasing) allele associated with reduced DIS3 expression. DIS3 encodes a catalytic subunit of the nuclear RNA exosome complex that mediates RNA processing and decay, and is mutated in several cancers. Chromosome conformation capture revealed a long range (570 kb) physical interaction between a sub-region of the risk locus, containing rs386772267, and a region ∼6 kb upstream of DIS3 Finally, repressor regulatory activity and allele-specific protein binding by transcription factors of the TCF/LEF family were observed for the risk-increasing allele of rs386772267, indicating that expression regulation at this risk locus may be influenced by the Wnt signaling pathway. In conclusion, we have identified a putative functional indel variant at chr13q22.1 that associates with decreased DIS3 expression in carriers of pancreatic cancer risk-increasing alleles, and could therefore affect nuclear RNA processing and/or decay.

Antibodies from donor B cells perpetuate cutaneous chronic graft-versus-host disease in mice.

Cutaneous sclerosis is one of the most common clinical manifestations of chronic graft-versus-host disease (cGVHD). Donor CD4(+) T and B cells play important roles in cGVHD pathogenesis, but the role of antibodies from donor B cells remains unclear. In the current studies, we generated immunoglobulin (Ig)H(µÎ³1) DBA/2 mice whose B cells have normal antigen-presentation and regulatory functions but cannot secrete antibodies. With a murine cGVHD model using DBA/2 donors and BALB/c recipients, we have shown that wild-type (WT) grafts induce persistent cGVHD with damage in the thymus, peripheral lymphoid organs, and skin, as well as cutaneous T helper 17 cell (Th17) infiltration. In contrast, IgH(µÎ³1) grafts induced only transient cGVHD with little damage in the thymus or peripheral lymph organs or with little cutaneous Th17 infiltration. Injections of IgG-containing sera from cGVHD recipients given WT grafts but not IgG-deficient sera from recipients given IgH(µÎ³1) grafts led to deposition of IgG in the thymus and skin, with resulting damage in the thymus and peripheral lymph organs, cutaneous Th17 infiltration, and perpetuation of cGVHD in recipients given IgH(µÎ³1) grafts. These results indicate that donor B-cell antibodies augment cutaneous cGVHD in part by damaging the thymus and increasing tissue infiltration of pathogenic Th17 cells.

Mechanical coupling of the multiple structural elements of the large-conductance mechanosensitive channel during expansion.

The prokaryotic mechanosensitive channel of large conductance (MscL) is a pressure-relief valve protecting the cell from lysing during acute osmotic downshock. When the membrane is stretched, MscL responds to the increase of membrane tension and opens a nonselective pore to about 30 Å wide, exhibiting a large unitary conductance of ∼ 3 nS. A fundamental step toward understanding the gating mechanism of MscL is to decipher the molecular details of the conformational changes accompanying channel opening. By applying fusion-protein strategy and controlling detergent composition, we have solved the structures of an archaeal MscL homolog from Methanosarcina acetivorans trapped in the closed and expanded intermediate states. The comparative analysis of these two new structures reveals significant conformational rearrangements in the different domains of MscL. The large changes observed in the tilt angles of the two transmembrane helices (TM1 and TM2) fit well with the helix-pivoting model derived from the earlier geometric analyses based on the previous structures. Meanwhile, the periplasmic loop region transforms from a folded structure, containing an ω-shaped loop and a short ß-hairpin, to an extended and partly disordered conformation during channel expansion. Moreover, a significant rotating and sliding of the N-terminal helix (N-helix) is coupled to the tilting movements of TM1 and TM2. The dynamic relationships between the N-helix and TM1/TM2 suggest that the N-helix serves as a membrane-anchored stopper that limits the tilts of TM1 and TM2 in the gating process. These results provide direct mechanistic insights into the highly coordinated movement of the different domains of the MscL channel when it expands.

MHC-mismatched mixed chimerism augments thymic regulatory T-cell production and prevents relapse of EAE in mice.

Multiple sclerosis (MS) is an autoimmune inflammatory disease of the central nervous system with demyelination, axon damage, and paralysis. Induction of mixed chimerism with allogeneic donors has been shown to not cause graft-versus-host disease (GVHD) in animal models and humans. We have reported that induction of MHC-mismatched mixed chimerism can cure autoimmunity in autoimmune NOD mice, but this approach has not yet been tested in animal models of MS, such as experimental autoimmune encephalomyelitis (EAE). Here, we report that MHC-mismatched mixed chimerism with C57BL/6 (H-2(b)) donor in SJL/J (H-2(s)) EAE recipients eliminates clinical symptoms and prevents relapse. This cure is demonstrated by not only disappearance of clinical signs but also reversal of autoimmunity; elimination of infiltrating T, B, and macrophage cells in the spinal cord; and regeneration of myelin sheath. The reversal of autoimmunity is associated with a marked reduction of autoreactivity of CD4(+) T cells and significant increase in the percentage of Foxp3(+) Treg among host-type CD4(+) T cells in the spleen and lymph nodes. The latter is associated with a marked reduction of the percentage of host-type CD4(+)CD8(+) thymocytes and an increase of Treg percentage among the CD4(+)CD8(+) and CD4(+)CD8(-) thymocytes. Thymectomy leads to loss of prevention of EAE relapse by induction of mixed chimerism, although there is a dramatic expansion of host-type Treg cells in the lymph nodes. These results indicate that induction of MHC-mismatched mixed chimerism can restore thymic negative selection of autoreactive CD4(+) T cells, augment production of Foxp3(+) Treg, and cure EAE.