Eugenol Possesses Colitis Protective Effects: Impacts on the TLR4/MyD88/NF-[Formula: see text]B Pathway, Intestinal Epithelial Barrier, and Macrophage Polarization.

Eugenol (EU) has been shown to ameliorate experimental colitis due to its anti-oxidant and anti-inflammatory bioactivities. In this study, DSS-induced acute colitis was established and applied to clarify the regulation efficacy of EU on intestinal barrier impairment and macrophage polarization imbalance along with the inflammatory response. Besides, the adjusting effect of EU on macrophages was further investigated in vitro. The results confirmed that EU intervention alleviated DSS-induced colitis through methods such as restraining weight loss and colonic shortening and decreasing DAI scores. Microscopic observation manifested that EU maintained the intestinal barrier integrity in line with the mucus barrier and tight junction protection. Furthermore, EU intervention significantly suppressed the activation of TLR4/MyD88/NF-[Formula: see text]B signaling pathways and pro-inflammatory cytokines gene expressions, while enhancing the expressions of anti-inflammatory cytokines. Simultaneously, WB and FCM analyses of the CD86 and CD206 showed that EU could regulate the DSS-induced macrophage polarization imbalance. Overall, our data further elucidated the mechanism of EU's defensive effect on experimental colitis, which is relevant to the protective efficacy of intestinal barriers, inhibition of oxidative stress and excessive inflammatory response, and reprogramming of macrophage polarization. Hence, this study may facilitate a better understanding of the protective action of the EU against UC.

Electroacupuncture regulates microglial polarization via inhibiting NF-κB/COX2 pathway following traumatic brain injury.

BACKGROUND: Neuroinflammation and oxidative stress are important pathological mechanisms following traumatic brain injury (TBI). The NF-κB/COX2 pathway regulates neuroinflammation and oxidative damage, while microglia also play an important role in neuroinflammation. Since NF-κB is involved in microglial polarization, targeting this pathway and microglial polarization is a critical component of TBI treatment. Currently, electroacupuncture (EA) is widely used to treat various symptoms after TBI, but the mechanisms of EA remain poorly understood. Additionally, the optimal frequency of EA remains unclear, which affects its efficacy. This study focuses on exploring the optimal frequency parameters of EA on TBI and investigating the underlying mechanisms of EA through NF-κB/COX2 pathway and microglial polarization. METHODS: The study was divided into two parts. In Experiment 1, 42 Sprague Dawley (SD) rats were induced and randomly divided into seven groups (n = 6). Except for the sham group, all rats underwent controlled cortical impact (CCI) to establish TBI model. Four EA groups (with different frequencies) and manual acupuncture (without current stimulation) received stimulation on the acupoints of Shuigou (GV26), Fengchi (GB20) and Neiguan (PC6) once a day for 7 days. The neurological function was assessed by modified Neurological Severity Scores (mNSS), and the rats' memory and learning were examined by the Morris water maze (MWM). SOD, MDA, and GSH-Px were detected to evaluate the levels of oxidative stress. The levels of IL-1ß, IL-6, and TNF-α were evaluated by Enzyme Linked Immunosorbent Assay (ELISA). Detection of the above indicators indicated a treatment group that exerted the strongest neuroprotection against TBI, we then conducted Experiment 2 using this screened acupuncture treatment to investigate the mechanism of acupuncture. 48 rats were randomly divided into four groups (n = 12): sham, TBI model, acupuncture and PDTC (NF-κB inhibitor). Evaluations of mNSS, MWM test, SOD, MDA, GSH-Px, IL-1ß, IL-6, TNF-α, and IL-10 were the same as in Experiment 1. Western blot was applied for detecting the expression levels of NF-κB, p-NF-κB, COX2, and Arg-1. TUNEL was used to examine neuronal apoptosis. Brain structure was observed by H&E. Iba-1, COX2, and Arg-1 were investigated by immunofluorescence staining. RESULTS: EA with frequency of 2/100 Hz markedly improved neuronal and cognitive function as compared to the other treatment groups. Moreover, it downregulated the expression of MDA, IL-6, IL-1ß, and TNF-α and upregulated the levels of SOD and GSH-Px. In addition, Both EA with 2/100 Hz and PDTC reduced the levels of p-NF-κB, COX2 and M1 markers (COX2, IL-6, IL-1ß, TNF-α) and increased the levels of M2 markers (Arg-1, IL-10). Moreover, they had similar effects on reducing inflammation, oxidative stress and apoptosis, and improving neuronal and cognitive function. CONCLUSIONS: The collective findings strongly suggest that EA with 2/100 Hz can improve neurologic function by suppressing neuroinflammation, oxidative stress and apoptosis. Additionally, we confirm that EA promotes microglial polarization towards the M2 phenotype through the suppression of NF-κB/COX2 pathway, thus exerting neuroprotective effects after TBI.

Electro-nape-acupuncture regulates the differentiation of microglia through PD-1/PD-L1 reducing secondary brain injury in acute phase intracerebral hemorrhage rats.

INTRODUCTION: This study aimed to investigate the effect of electro-nape-acupuncture (ENA) on the differentiation of microglia and the secondary brain injury in rats with acute-phase intracerebral hemorrhage (ICH) through the programmed cell death protein-1/ligand-1 (PD-1/PD-L1) pathway. METHODS: A total of 27 male Sprague-Dawley rats were randomly divided into three groups: sham group, ICH group, and ENA group. The autologous blood infusion intracerebral hemorrhage model was used to study the effects of ENA by administering electroacupuncture at GB20 (Fengchi) and Jiaji (EX-B2) acupoints on 24 h after the modeling, once per day for 3 days. The neurological function damage, hematoma lesion, and inflammatory cell infiltration were measured by the beam walking test and hematoxylin-eosin staining. The expression of PD-1, PD-L1, CD86, CD206, and related cytokines around the hematoma was measured by western blot, quantitative reverse transcription polymerase chain reaction, and immunofluorescence. RESULTS: The ICH group had significant neurological deficits (p < .001), hematoma lesions, and inflammatory cell infiltration. The levels of CD86 protein, inflammatory factors tumor necrosis factors (TNF)-α, interleukin (IL)-1ß, and IL-6 were increased (p < .001), while CD206 protein was reduced (p < .01), and the number of CD86+ /CD11b+ cells was also increased (p < .001) compared to the sham group. However, after ENA intervention, there was a significant reduction in neurological function damage (p < .05), infiltration of inflammatory cells, and the expression levels of CD86+ /CD11b+ cells (p < .05), resulting in the increased expression of PD-1 protein and differentiation of M2 phenotype significantly (p < .001). CONCLUSION: The study concludes that ENA could reduce neurological function damage, inhibit the expression of pro-inflammatory cytokines, and improve the infiltration of inflammatory cells to improve secondary brain injury in acute-phase intracerebral hemorrhage rats. These effects could be related to the increased expression of PD-1 around the lesion, promoting the differentiation of microglia from M1 to M2 phenotype.

Rapamycin Alleviates 2,4,6-Trinitrobenzene Sulfonic Acid-Induced Colitis through Autophagy Induction and NF-κB Pathway Inhibition in Mice.

Background: Recent genetic studies indicated that variants of autophagy genes were associated with the predisposition of Crohn's disease (CD). The autophagy deficiency may affect the innate and adaptive immunity, which is related to persistent and excessive inflammation of the bowel. However, it remains unclear how autophagy modulates the expression of immune response regulator NF-κB and proinflammatory cytokine TNF-α in CD. Aim: We aimed to investigate the role of rapamycin on the expression of NF-κB p65 and TNF-α in 2, 4, 6-trinitrobenzene sulfonic acid (TNBS)-induced mouse colitis and lipopolysaccharide (LPS)-induced HT-29 cells. Methods: TNBS-induced colitis mice were treated with saline or rapamycin, and the disease activity index (DAI) and histological scores of colonic mucosa were evaluated. The expressions of p65, ATG16L1 and LC3 were detected by western blot and immunohistochemistry staining. The monodansylcadaverine (MDC) staining and transmission electron microscopy were developed to study the autophagy in LPS-induced HT-29 cells. Expression of TNF-α from colon tissue and HT-29 cells were detected by ELISA. The expressions of p65, ATG16L1 and LC3 in active CD patients were also investigated. Results: Significantly more autophagosomes were observed in rapamycin-treated cells than in controls. Rapamycin remarkably upregulated the expression of ATG16L1 and LC3II, inhibited p65 nucleus translocation and secretion of TNF-α both in vivo and in vitro. The expression of both ATG16L1 and LC3II increased in mild to moderate CD specimens, while no significant difference was noted between severe CD and normal controls. The expression of p65 increased notably in severe CD compared to those in mild to moderate patients. Conclusions: In LPS-treated HT-29 cells and TNBS-induced colitis, p65 is overexpressed, which results in exaggerated secretion of TNF-α and induce or worsen the inflammation in the bowel. Rapamycin protects against colitis through induction of autophagy, thus inhibiting the activation of NF-κB pathway and secretion of TNF-α.

Isolated hepatic tuberculosis associated with portal vein thrombosis and hepatitis B virus coinfection: A case report and review of the literature.

BACKGROUND: While tuberculosis (TB) itself is a common disease, isolated TB of the liver is a rare entity. Tubercular involvement of the liver is more commonly a part of a disseminated disease of the hepatic parenchyma. In contrast, isolated hepatic TB spread through the portal vein from the gastrointestinal tract is seldom encountered in clinical practice, with only a few sporadic cases and short series available in the current literature. Vascular complications, such as portal vein thrombosis (PVT), have rarely been reported previously. CASE SUMMARY: A 22-year-old man was hospitalized with complaints of a 3-mo history of fever and weight loss of approximately 10 kg. He had a 10-year hepatitis B virus (HBV) infection in his medical history. Contrast-enhanced computed tomography (CECT) confirmed hepatosplenomegaly, with hypodensity of the right lobe of the liver and 2.1 cm thrombosis of the right branch of the portal vein. A liver biopsy showed epithelioid granulomas with a background of caseating necrosis. Ziehl-Nelson staining showed acid-fast bacilli within the granulomas. The patient was diagnosed with isolated hepatic TB with PVT. Anti-TB therapy (ATT), including isoniazid, rifapentine, ethambutol, and pyrazinamide, was administered. Along with ATT, the patient was treated with entecavir as an antiviral medication against HBV and dabigatran as an anticoagulant. He remained asymptomatic, and follow-up sonography of the abdomen at 4 mo showed complete resolution of the PVT. CONCLUSION: Upon diagnosis of hepatic TB associated with PVT and HBV coinfection, ATT and anticoagulants should be initiated to prevent subsequent portal hypertension. Antiviral therapy against HBV should also be administered to prevent severe hepatic injury.

Correlation between serum 25-(OH)D3 level and immune imbalance of Th1/Th2 cytokines in patients with Hashimoto's thyroiditis and its effect on autophagy of human Hashimoto thyroid cells.

The study aimed to determine the relationship between serum 25-(OH)D3 and Th1/Th2 cytokine immune imbalance, and the effect of 25-(OH)D3 on the autophagy of human Hashimoto thyroid cells. Western blot analysis was used to detect the expression levels of microtubule-associated protein 1 light chain 3 (LC3) and autophagy-associated protein mammalian target protein of rapamycin (mTOR) in thyroid tissues of 20 Hashimoto's thyroiditis (HT) patients and normal tissues of 20 benign thyroid adenomas. Nthy-ori3-1 cells (normal cells of human thyroid follicular epithelium) were treated with different concentrations of 25-(OH)D3 for 24 h. The expression of LC3, mTOR and caspase-3 protein in the cells was detected by western blot analysis. The apoptosis and proliferation levels were detected by flow cytometry and MTT assay, respectively. The levels of FT3, FT4 and IL-10 in the HT group were lower than those in the healthy control group. The serum levels of 25-(OH)D3, TPOAb and TGAb in the HT group were lower than those in the healthy control group. Serum 25-(OH)D3 level in the HT group was negatively correlated with IL-2 and IFN-γ, and positively correlated with IL-4. In Hashimoto's thyroiditis tissues, the expression of mTOR was higher while the expression of LC3B-II was lower than that of normal thyroid tissue. With the increase in 25-(OH)D3 concentration, the expression level of mTOR increased, the expression level of LC3B-II decreased and the apoptosis rate was significantly increased. The cell proliferation rate decreased with the increase in 25-(OH)D3 concentration. The serum 25-(OH)D3 level in HT hypothyroidism patients was significantly lower than that of the control group. Thus, 25-(OH)D3 may be involved in the disease progression by upregulating the levels of Th1 cytokines and downregulating the levels of Th2 cytokines. 25-(OH)D3 can inhibit autophagy of thyroid cells, induce apoptosis and participate in the pathogenesis of Hashimoto's thyroiditis.

Transplantation of human matrix metalloproteinase-1 gene-modified bone marrow-derived mesenchymal stem cell attenuates CCL4-induced liver fibrosis in rats.

It has been reported that bone marrow-derived mesenchymal stem cells (BMSCs) alleviated liver fibrosis. We investigated whether BMSCs transfected with human matrix metalloproteinase 1 (BMSCs/MMP1) would improve their therapeutic effect in liver fibrosis induced by carbon tetrachloride (CCl4) in rats. BMSCs were transfected with an adenovirus carrying enhanced green fluorescence protein (GFP) and human MMP1 gene. BMSCs or BMSCs/MMP1 were directly injected into fibrotic rats via the tail vein. GFP-labeled cells appeared in the fibrotic liver after BMSC transplantation. The expression of BMSCs/MMP1 elevated levels of MMP1 in vitro. Although BMSC administration reduced liver fibrosis, transplantation of BMSCs/MMP1 enhanced the reduction of liver fibrosis to a higher level. Treatment with BMSCs/MMP1 not only decreased collagen content but also suppressed activation of hepatic stellate cells (HSCs) in fibrotic liver, which led to subsequent improvement of both liver injury and fibrosis. Treatment with BMSCs/MMP1 resulted in an improved therapeutic effect compared with BMSCs alone, which is probably because of the sustainably expressed MMP1 level in the liver. BMSCs/MMP1 transplantation not only improved biochemical parameters but also attenuated progression of liver fibrosis, suggesting that BMSCs may be a potential cell source in preventing liver fibrosis and MMP1 gene may enhance the anti-fibrotic effect of BMSCs.

Pancreatic T/histiocyte-rich large B-cell lymphoma: A case report and review of literature.

Primary pancreatic lymphoma (PPL) is an extremely rare form of extranodal malignant lymphoma. The most common histological subtype of PPL is diffuse large B cell lymphoma (DLBCL). In rare cases, PPL can also present as follicular lymphoma, small lymphocytic lymphoma, and T cell lymphoma either of non-Hodgkin's lymphoma or of Hodgkin's lymphoma. T-cell/histiocyte-rich large B-cell lymphoma (T/HRBCL) is an uncommon morphologic variant of DLBCL with aggressive clinical course, it is predominantly a nodal disease, but extranodal sites such as bone marrow, liver, and spleen can be involved. Pancreatic involvement of T/HRBCL was not presented before. Herein, we report a 48-year-old male who was hospitalized with complaints of jaundice, dark brown urine, pale stools, and nausea. The radiological evaluation revealed a pancreatic head mass and, following operative biopsy, the tumor was diagnosed as T/HRBCL. The patient achieved remission after six cycles of CHOP chemotherapy. Therefore, T/HRBCL can be treated similarly to the stage-matched DLBCL and both of them get equivalent outcomes after chemotherapy.

[Therapeutic effect of BMSCs with over-expressed MMP1 on liver fibrosis].

OBJECTIVE: To investigate the function of bone marrow mesenchymal stem cells (BMSCs) with over-expressed matrix metalloproteinase 1 (MMP1) on liver fibrosis. METHODS: Fifty SD male rats were randomly divided into 4 groups: recombinant adenovirus Adhuman MMP-1(hMMP-1)-enhanced green fluorescent protein (EGFP) transfected BMSCs group (Group A, n=10), Ad-EGFP transfected BMSCs group (Group B, n=10), liver fibrosis group (Group C, n=15), and a normal group (Group D, n=15). The liver fibrosis model was formed by subcutaneous injection of the mixed liquor of carbon tetrachloride (CCL4) and vegetable oil. After 10 weeks, the model of liver fibrosis was formed. Group A and B were administered the transfected BMSCs via the tail veins, while Group C and D were administered normal saline. After 3 weeks, the rats were sacrificed. The body weight, liver weight, liver function, liver fibrosis indexes and liver pathological changes were tested. RESULTS: Compared with the control group, the rats administered BMSCs with over-expressed MMP1 showed a significant improvement in the body weight, liver weight and plasma albumin (ALB) (P<0.05), and a significant reduction in the plasma alanine aminotransferase, total bilirubin, hyaluronic acid, laminin and procollagen III (P<0.05). Hematoxylin-eosin staining confirmed that the degree of liver fibrosis was significantly ameliorated under average visual fields (P<0.05). CONCLUSION: The repair ability of BMSCs on liver fibrosis can be enhanced by over-expression of hMMP-1.

[Recombinant adenovirus Ad-human matrix metalloproteinase 1 transfecting bone marrow mesenchymal stem cells of rats in vitro].

OBJECTIVE: To transfect bone marrow mesenchymal stem cells (BMSCs) of rats by recombinant adenovirus Ad-human matrix metalloproteinase 1 (hMMP-1) in vitro so as to lay the experimental foundation for the treatment of liver fibrosis with a combination of BMSCs and hMMP-1 gene transplantation. METHODS: BMSCs were isolated from bone marrow of 2-3 weeks old Sprague Dawley rats by whole bone marrow adherence method and identified, then transfected by recombinant adenovirus Ad-hMMP-1 carrying enhanced green fluorescent protein (EGFP) marker in vitro. The green fluorescent expression was observed by fluorescence microscope and the transfection efficiency was detected by flow cytometry to determine the optimum multiplicity of infection (MOI). BMSCs at passage 3 were divided into 3 groups: untransfected BMSCs group (group A), Ad-EGFP transfected BMSCs group (group B), and Ad-hMMP-1-EGFP transfected BMSCs group (group C); the gene and intracellular protein of hMMP-1 were detected by RT-PCR and Western blot; the ELISA assay was used to detect the supernatant protein expression, and the hMMP-1 activity was measured by fluorescent quantification kit. RESULTS: The green fluorescent was observed in BMSCs transfected by recombinant adenovirus at 24 hours after transfection; the fluorescence intensity was highest at 72 hours; and the optimum MOI was 200. The cells of 3 groups entered the logarithmic growth phase on the 3rd day and reached plateau phase on the 6th day by MTT assay; no significant difference was found in the cell proliferation rate among 3 groups (P > 0.05). RT-PCR, Western blot, and ELISA assay showed high expressions of the hMMP-1 gene and protein in group C, but no expression in groups A and B. The hMMP-1 activity was 1.24 nmol/(mg.min) in group C, but hMMP-1 activity was not detectable in groups A and B. CONCLUSION: The exogenous hMMP-1 gene is successfully transfected into BMSCs of rats via recombinant adenovirus and can highly express, which lays the experimental foundation for the treatment of liver fibrosis with a combination of BMSCs and hMMP-1 gene transplantation.

A case of rapid growing colonic NK/T cell lymphoma complicated by Crohn's disease.

A 37-year-old man developed abdominal pain and bloody diarrhea 11 months before admission. The colonoscopy revealed multifocal ulcers in the colon. Histology showed active chronic inflammation. Although anti-tuberculosis medication was effective, his symptoms repeated 2 months later. The subsequent colonoscopy revealed more extensive irregular ulcers than before, and he was clinically suspected with intestinal malignant lymphoma. He underwent subtotal colectomy and was histologically suggested Crohn's disease, then 5-aminosalicylic and a combination of prednisone and azathioprine were administered in succession postoperatively, but they achieved minimal relief of symptoms for a period of 7 months. The third colonoscopy showed a large irregular ulcer in the ileocolon stomas, and primary colonic NK/T cell lymphoma was diagnosed through histological and immunophenotypic studies. Malignant lymphoma should be taken into consideration when clinically diagnosed Crohn's disease was refractory to medication or when its clinical course became aggressive.

Primary intestinal NK/T cell lymphoma: a clinicopathologic study of 25 Chinese cases.

BACKGROUND: Primary intestinal NK/T cell lymphoma is extremely rare and early diagnosis is frequently difficult. The aim of this study is to investigate the clinicopathological findings, immunophenotype, and T cell receptor (TCR) γ gene rearrangement of primary intestinal NK/T cell lymphomas in 25 Chinese cases. METHODS: Clinical data of the 25 cases were analyzed. Immunohistochemistry for immunophenotype, in situ hybridization for EBER, and polymerase chain reaction for TCR γ gene rearrangement were investigated. Survival curves according to clinical characteristics were analyzed. RESULTS: The median age was 33 years and the median survival was 7 months. The common symptoms consisted of abdominal pain, fever, marasmus, diarrhea, and hematochezia. Endoscopically, the tumors were mainly featured by focal, multifocal or diffuse irregular ulcers, which most frequently emerged in the ascending colon. Histologically, the tumors were characterized by the proliferation of pleomorphic atypical lymphoid cells (ALCs), necrosis, lympho-epithelial lesions, and mixed inflammatory infiltration. The positive frequency of CD3ε was 88.2%, of CD56 was 84%, granzyme B was 90%, and EBER was 84.2%. A total of 12 out of 14 cases (85.7%) highly expressed Ki67. The negative prognostic factors for survival were Ann Arbor stage IIIE or IVE (P = 0.039) and more than one extranodal site of disease (P = 0.019). CONCLUSION: Primary intestinal NK/T cell lymphomas most frequently favor young people and have a poor prognosis. Due to the nonspecific clinical and endoscopic findings, it is difficult to distinguish intestinal NK/T cell lymphomas from inflammatory and infectious disorders. Histopathology, immunophenotype, and DNA study play key roles in differential diagnosis.

Gut-liver axis plays a role in hepatocarcinogenesis of patients with Crohn's disease.

The development of hepatocellular carcinoma (HCC) is attributed to several factors, including chronic viral infection, alcohol consumption, exposure to aflatoxin B1 and metabolic disorders. Several recent reports have shown that HCC can occur in patients with long-standing Crohn's disease (CD) in the absence of other underlying high-risk liver diseases. There may be an association between CD and hepatocarcinogenesis, however, the precise mechanism for this requires further investigations.

An experimental study of extracellular signal-regulated kinase and its interventional treatments in hepatic fibrosis.

BACKGROUND: The pathogenesis of hepatic fibrosis and cirrhosis is still not fully understood. The extracellular signal-regulated kinase (ERK) pathway is involved in the regulation of cell proliferation and differentiation. The aim of this study was to investigate the effects of PD98059, a specific inhibitor of ERK, on the cell cycle, cell proliferation, secretion of type I collagen and expression of cyclin D1 mRNA, CDK4 mRNA and transforming growth factor-beta1 (TGF-beta1) mRNA in rat hepatic stellate cells (HSCs) stimulated by acetaldehyde. METHODS: Rat HSCs stimulated by acetaldehyde were incubated with PD98059 at different concentrations. The cell cycle was analysed by flow cytometry. Cell proliferation was assessed by the methyl thiazolyl tetrazolium colorimetric assay. The mRNA expression of cyclin D1, CDK4 and TGF-beta1 was examined using the reverse transcriptase-polymerase chain reaction. Type I collagen in the culture medium was detected by enzyme-linked immunosorbent assay. RESULTS: 20, 50 and 100 micromol/L PD98059 significantly inhibited the proliferation and provoked a G0/G1-phase arrest of acetaldehyde-induced HSCs in a dose-dependent manner. The secretion of type I collagen and the expression of cyclin D1, CDK4 and TGF-beta1 mRNA in acetaldehyde-induced HSCs were markedly inhibited by 50 and 100 micromol/L PD98059, respectively. CONCLUSIONS: The ERK pathway regulates the cell proliferation, secretion of type I collagen and the expression of TGF-beta1 mRNA in rat HSCs stimulated by acetaldehyde, which is likely related to its regulative effect on the cell cycle.