To evaluate the protective effects of dexmedetomidine on the colon and spinal cord of rats with functional chronic visceral pain and its regulatory mechanism on extracellular signal-regulated kinase 1-cyclic adenosine monophosphate response element-binding protein signaling pathway. A rat model of irritable bowel syndrome was established by colorectal distention stimulation. Animals were categorized into normal group, model group, experimental group and control group. No treatment was given in normal group, while colorectal distention stimulation was utilized in model group, experimental group and control group. After successful modeling, the animals in experimental group were injected intraperitoneally with 5 μg/kg of dexmedetomidine hydrochloride injection daily, those in control group were injected intraperitoneally with 5 μg/kg pinaverium bromide daily and those in normal group and model group were injected intraperitoneally daily with the same volume of normal saline, for 14 consecutive days. Abdominal withdrawal reflex score and pain threshold of rats were measured. Cell apoptosis in the spinal cord of rats was determined by terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling staining. The messenger ribonucleic acid expressions of extracellular signal-regulated kinase 1 and cyclic adenosine monophosphate response element-binding protein in the spinal cord were detected by realtime quantitative polymerase chain reaction and phosphorylated-extracellular signal-regulated kinase 1 and phosphorylated-cyclic adenosine monophosphate response element-binding protein expressions in the spinal cord of rats were tested by Western blotting. Compared with normal group, abdominal withdrawal reflex score, cell apoptosis rate, messenger ribonucleic acid expressions of extracellular signal-regulated kinase 1 and cyclic adenosine monophosphate response element-binding protein, phosphorylatedextracellular signal-regulated kinase 1 and phosphorylated-cyclic adenosine monophosphate response element-binding protein expressions were significantly higher (p<0.05) and pain threshold value was significantly lower (p<0.05) in model group, experimental group and control group at 20 mmHg, 40 mmHg, 60 mmHg and 80 mmHg pressures. Abdominal withdrawal reflex score, cell apoptosis rate, messenger ribonucleic acid expressions of extracellular signal-regulated kinase 1 and cyclic adenosine monophosphate response element-binding protein, phosphorylated-extracellular signal-regulated kinase 1 and phosphorylated-cyclic adenosine monophosphate response element-binding protein expressions were significantly lower (p<0.05) and pain threshold value was significantly higher (p<0.05) in experimental group and control group than those in model group at 20 mmHg, 40 mmHg, 60 mmHg and 80 mmHg pressures. Dexmedetomidine can prominently ameliorate the clinical symptoms of rats with functional chronic visceral pain and may protect the colon and spinal cord by inhibiting the extracellular signalregulated kinase 1/cyclic adenosine monophosphate response element-binding protein signaling pathway.

IL-33 promotes type 2 immunity, epithelial repair, and tissue fibrosis by activating group 2 innate lymphoid cells (ILC2). ILC2 lack all known surface markers of mature T, B, NK, and myeloid cell lineages (Linneg), express the IL-33 receptor ST2, and release type 2 cytokines which contribute to cholangiocyte proliferation and activation of hepatic stellate cells. This pathway results in massive proliferation of the extrahepatic bile duct (EHBD) but also exacerbates liver fibrosis, suggesting that there may be tissue-specific subpopulations of IL-33-induced ILC. To determine the tissue-specific subsets of ILC in the hepatobiliary system, we analyzed CD45+Linneg mononuclear cells from IL-33 treated adult Balb/c mouse liver or EHBD by single cell RNA sequencing. Principal component analysis identified 6 major CD45+Linneg cell classes, two of which were restricted to the EHBD. One of these classes, biliary immature myeloid (BIM) cells, was predicted to interact with ILC2 by a network of shared receptor-ligand pairs. BIM highly expressed Gp49 and ST2 receptors on the cell surface while lacking surface expression of markers for mature myeloid cells. In conclusion, single cell RNA sequencing identified IL-33 responsive cell groups regionally confined to the liver or extrahepatic bile duct, including a novel population of CD45+Linneg Gp49-expressing mononuclear cells.

Correlation of serum vitamin E content with insulin resistance and oxidative stress response in patients with type 2 diabetes mellitus Jun Li, Ling Han, Li Dong, Shu-Lei Liu 1. Department of Clinical Laboratory Medicine, Xining Second People’s Hospital in Qinghai Province, Xining City, Qinghai Province, 810003 2. Department of Obstetrics, Xining Second People’s Hospital in Qinghai Province, Xining City, Qinghai Province, 810003 3. Department of Clinical Laboratory Medicine, the Fourth People’s Hospital of Qinghai Province, Xining City, Qinghai Province, 810001 Journal of Hainan Medical University

Injury to the biliary epithelium triggers inflammation and fibrosis, which can result in severe liver diseases and may progress to malignancy. Development of a type 1 immune response has been linked to biliary injury pathogenesis; however, a subset of patients with biliary atresia, the most common childhood cholangiopathy, exhibit increased levels of Th2-promoting cytokines. The relationship among different inflammatory drivers, epithelial repair, and carcinogenesis remains unclear. Here, we determined that the Th2-activating cytokine IL-33 is elevated in biliary atresia patient serum and in the livers and bile ducts of mice with experimental biliary atresia. Administration of IL-33 to WT mice markedly increased cholangiocyte proliferation and promoted sustained cell growth, resulting in dramatic and rapid enlargement of extrahepatic bile ducts. The IL-33-dependent proliferative response was mediated by an increase in the number of type 2 innate lymphoid cells (ILC2s), which released high levels of IL-13 that in turn promoted cholangiocyte hyperplasia. Induction of the IL-33/ILC2/IL-13 circuit in a murine biliary injury model promoted epithelial repair; however, induction of this circuit in mice with constitutive activation of AKT and YAP in bile ducts induced cholangiocarcinoma with liver metastases. These findings reveal that IL-33 mediates epithelial proliferation and suggest that activation of IL-33/ILC2/IL-13 may improve biliary repair and disruption of the circuit may block progression of carcinogenesis.

Abstract Id3-/- mice have been shown to develop a T cell-dependent autoimmune-like sjogren’s syndrome(SS). The dysregulated differentiation between Foxp3+ Tregs and Th17+ T cells in Id3-/- CD4+ T cells promoted us to study whether the autoimmune-like SS in Id3-/- mice was associated with the abnormal expression of Th17 cells in the target organs. We focused on the salivary glands and lachrymal glands, as they are main target tissues involving in SS. 50% of Id3-/- started to show proinflammatory CD4+ infiltrating cells in the salivary glands. Significantly higher frequency of Th1 and Th17 cells were observed in Id3-/- salivary glands, and higher frequency of Th17 and lower frequency of Tregs were observed in Id3-/- spleen. Notably, 26% mice Id3-/- mice developed gd T cell lymphoma. The gd T lymphoma cells were characterized by dysregulated IL-17 production and high expression of an oncogene, c-myc. Adoptive transfer of Id3-/- gd T lymphoma cells to Rag1-/- mice resulted in gd T lymphoma, suggesting that Id3 may regulates gd T cell tumor development.

Biliary atresia (BA) is a destructive cholangiopathy of childhood in which Th1 immunity has been mechanistically linked to the bile duct inflammation and obstruction that culminate in liver injury.Based on reports of decreased Th1 cytokines in some patients and the development of BA in mice lacking CD4 + T cells, we hypothesized that Th1-independent mechanisms can also activate effector cells and induce BA.Here, we tested this hypothesis using Stat1 -/-mice, which lack the ability to mount Th1 immune responses.Infection of Stat1 -/- mice with rhesus rotavirus type A (RRV) on postnatal day 1 induced a prominent Th2 response, duct epithelial injury and obstruction within 7 days, and atresia shortly thereafter.A high degree of phosphorylation of the Th2 transcription factor Stat6 was observed; however, concurrent inactivation of Stat1 and Stat6 in mice did not prevent BA after RRV infection.In contrast, depletion of macrophages or combined loss of Il13 and Stat1 reduced tissue infiltration by lymphocytes and myeloid cells, maintained epithelial integrity, and prevented duct obstruction.In concordance with our mouse model, humans at the time of BA diagnosis exhibited differential hepatic expression of Th2 genes and serum Th2 cytokines.These findings demonstrate compatibility between Th2 commitment and the pathogenesis of BA, and suggest that patient subgrouping in future clinical trials should account for differences in Th2 status.

Human γδ T-cell lymphoma is a rare clinicopathologic entity with aggressive course and poor prognosis. The etiology and pathogenesis of γδ T-cell lymphoma is unknown. We show here that mice with deficiency in inhibitory helix-loop-helix protein Id3 (Id3(-/-)) developed γδ T-cell lymphoma that resembled human γδ T-cell lymphoma. The Id3(-/-) mice with lymphoma showed splenomegaly, hepatomegaly, and lymphadenopathy with involvement of bone marrow, thymus, kidney, and lungs between 6 and 15 months of age. Phenotypic analysis revealed that lymphomatous cells were cluster of differentiation (CD)3(+), γδ T-cell receptor (TCR)(+), and αβ TCR(-), and expressed CD8(+)CD4(-), CD4(+)CD8(-), or a mixture of the two. Id3(-/-) γδ T-cell lymphoma used predominantly Vγ1.1, some Vγ3, yet no Vγ2 TCR, and some showed increased levels of the oncogene c-Myc. Strikingly, adoptive transfer of the γδ T-cell lymphoma into syngeneic Rag1(-/-) mice resulted in aggressive γδ T-cell lymphoma, identical to the Id3(-/-) donor. Thus, our data demonstrate that Id3 regulates the development of γδ T-cell lymphoma in mice, raising a possibility of Id3 gene mutation in human γδ T-cell lymphoma. Our model will provide a tool for studying the molecular mechanisms and development of human γδ T-cell lymphoma.

Human (cid:1)(cid:2) T-cell lymphoma is a rare clinicopathologic entity with aggressive course and poor prognosis. The etiology and pathogenesis of (cid:1)(cid:2) T-cell lymphoma is unknown. We show here that mice with deficiency in inhibitory helix-loop-helix protein Id3 (Id3 (cid:3) / (cid:3) ) developed (cid:1)(cid:2) T-cell lymphoma that resembled human (cid:1)(cid:2) T-cell lymphoma. The Id3 (cid:3) / (cid:3) mice with lymphomashowedsplenomegaly,hepato-megaly, and lymphadenopathy with involvement of bone marrow, thymus, kid-ney, and lungs between 6 and 15 months of age. Phenotypic analysis revealed that lymphomatous cells were cluster of differentiation (CD)3 (cid:4) , (cid:1)(cid:2) T-cell receptor (TCR) (cid:4) , and (cid:5)(cid:6) TCR (cid:3) , and expressed CD8 (cid:4) CD4 (cid:3) , CD4 (cid:4) CD8 (cid:3) , or a mixture of the two. Id3 (cid:3) / (cid:3) (cid:1)(cid:2) T-cell lymphoma used predominantly V (cid:1) 1.1, some V (cid:1) 3, yet no V (cid:1) 2 TCR, and some showed increased levels of the oncogene c-Myc. Strikingly, adoptive transfer of the (cid:1)(cid:2) T-cell lymphoma into syngeneic Rag1 (cid:3) / (cid:3) mice re-sulted in aggressive (cid:1)(cid:2) T-cell lymphoma, identical to the Id3 (cid:3) / (cid:3) donor. Thus, our data demonstrate that Id3 regulates the development of (cid:1)(cid:2) T-cell lymphoma in mice, raising a possibility of Id3 gene mutation in human (cid:1)(cid:2) T-cell lymphoma. Our model will provide a tool for studying the molecular mechanisms and development of human (cid:1)(cid:2) T-cell lymphoma.

The molecular mechanisms that direct transcription of the gene encoding the transcription factor Foxp3 in CD4(+) T cells remain ill-defined. We show here that deletion of the DNA-binding inhibitor Id3 resulted in the defective generation of Foxp3(+) regulatory T cells (T(reg) cells). We identify two transforming growth factor-β1 (TGF-β1)-dependent mechanisms that were vital for activation of Foxp3 transcription and were defective in Id3(-/-) CD4(+) T cells. Enhanced binding of the transcription factor E2A to the Foxp3 promoter promoted Foxp3 transcription. Id3 was required for relief of inhibition by the transcription factor GATA-3 at the Foxp3 promoter. Furthermore, Id3(-/-) T cells showed greater differentiation into the T(H)17 subset of helper T cells in vitro and in a mouse asthma model. Therefore, a network of factors acts in a TGF-β-dependent manner to control Foxp3 expression and inhibit the development of T(H)17 cells.

The molecular mechanisms directing the development of 'natural' CD4 + CD25 + Foxp3 + regulatory T cells (T reg cells) in the thymus are not thoroughly understood. We show here that conditional deletion of transforming growth factor-β receptor I (TβRI) in T cells blocked the appearance of CD4 + CD25 + Foxp3 + thymocytes at postnatal days 3–5. Paradoxically, however, beginning 1 week after birth, the same TβRI-mutant mice showed accelerated expansion of thymic CD4 + CD25 + Foxp3 + populations. This rapid recovery of Foxp3 + thymocytes was attributable mainly to overproduction of and heightened responsiveness to interleukin 2, as genetic ablation of interleukin 2 in TβRI-mutant mice resulted in a complete absence of CD4 + CD25 + Foxp3 + cells from the thymus and periphery. Thus, transforming growth factor-β signaling is critical to the thymic development of natural CD4 + CD25 + Foxp3 + T reg cells.

Background: CD4(+)CD25(+) T regulatory cells (Tregs) play an important role in regulating immune responses, and in influencing human immune diseases such as HIV infection. It has been shown that human CD4(+)CD25(+) Tregs can be induced in vitro by TCR stimulation of CD4(+)CD25(-) T cells. However, the mechanism remains elusive, and intriguingly, similar treatment of murine CD4(+)CD25(-) cells did not induce CD4(+)CD25(+)Foxp3(+) Tregs unless exogenous TGF-beta was added during stimulation. Thus, we investigated the possible role of TGF-beta in the induction of human Tregs by TCR engagement. We also explored the effects of TGF-beta on HIV-1 infection mediated induction of human Tregs since recent evidence has suggested that HIV-1 infection may also impact the generation of Tregs in infected patients. Results: We show here that endogenous TGF-beta is key to TCR induction of Foxp3 in human CD4(+)CD25(-) T cells. These events involve, first, the production of TGF-beta by TCR and CD28 stimulation and the activation of latent TGF-beta by reactive oxygen species generated from the activated T cells. Biologically active TGF-beta then engages in the induction of Foxp3. Neutralization of active TGF-beta with anti-TGF-beta antibody or elimination of ROS with MnTBAP abrogated Foxp3 expression. HIV-1 infection enhanced Foxp3 expression in activated CD4(+)CD25(-) T cells; which was also abrogated by blockade of endogenous TGF-beta. Conclusion: Several conclusions can be drawn from this work: (1) TCR and CD28-induced Foxp3 expression is a late event following TCR stimulation; (2) TGF-beta serves as a link in Foxp3 induction in human CD4(+)CD25(-) T cells following TCR stimulation, which induces not only latent, but also active TGF-beta; (3) the activation of TGF-beta requires reactive oxygen species; (4) HIV infection results in an increase in Foxp3 expression in TCR-activated CD25(-) T cells, which is also associated with TGF-beta. Taken together, our findings reinforce a definitive role of TGF-beta not only in the generation of Tregs with respect to normal immune responses, but also is critical in immune diseases such as HIV-1 infection.

Three major mucosal systems exist in the body, the oral-gastrointestinal, the respiratory and the genitourinary systems. In particular, the gastrointestinal (GI) tract contains the largest mucosal surface in the body and is the major port of entry for foreign antigens. Therefore, the gut immune system has to differentiate to tolerate dietary antigens and expel infectious and harmful pathogens. During the complex but well-orchestrated immune responses in the mucosal system, T cells play a pivotal role in both immunity and tolerance. Of many T cell subpopulations, CD4(+)CD25(+) T regulatory cells (Tregs) are instrumental in regulation of immune responses in mucosea. Among the multitude of cytokines and factors that are produced in the gut, Transforming Growth Factor-beta (TGF-beta) is probably the most important one in influencing mucosal T cell responses. The interaction and mutual regulation between TGF-beta and CD4(+)CD25(+) Tregs may be the key in maintaining the balance between T cell immunity and tolerance in mucosal system. In this article, we attempt to discuss both beneficial and detrimental effects of TGF-beta and Tregs on oral tolerance, mucosal inflammation and autoimmunity, colon cancer and HIV infection in the gut.