Mucosal Immunity Related to CD8+ T Lymphocytes in Children with Helicobacter pylori Gastritis.

Purpose: We investigated the role of CD8+T cells as host immune factors in pediatric patients with Helicobacter pylori gastritis. Methods: Gastric mucosal tissue and blood samples were collected from 39 children, including 11 children with H. pylori infection and 28 children as controls. Anti-CD8 and anti-T-bet antibodies were used for immunohistochemistry of the gastric mucosa. For the cell surface and intracellular staining, peripheral blood mononuclear cells were stained with anti-IL7Rα, anti-CX3CR1, anti-CD8, anti-T-bet, and anti-IFN-γ antibodies. Cytokines of sera such as tumor necrosis factor alpha (TNF-α) and CX3CL1 were analyzed using enzyme- linked immunosorbent assay (ELISA). Results: In the immunohistochemistry of gastric mucosa, the frequency of CD8+ and T-bet+ T cells cells was higher in the H. pylori-positive group than in the control group (26.9± 7.8% vs. 16.9±3.3%, p<0.001; 5.0±2.5% vs. 2.2±0.7%, p=0.001). Between the control and H. pylori-positive groups, the frequency of IL-7RαlowCX3CR1+ CD8+ and T-bet+ INF-γ+ CD8+ T cells were not significantly different between surface and intracellular staining, respectively (40.4±24.0% vs. 38.2±17.8%, p=0.914; 40.4±24.0% vs. 38.2±17.8%, p=0.914). In the ELISA, no significant differences in TNF-α and CX3CL1 concentrations were observed between the control and H. pylori-positive groups (34.3±12.1 pg/mL vs. 47.0±22.6 pg/mL, p=0.114/0.5± 0.1 pg/mL vs. 0.5±0.1 pg/mL, p=0.188). Conclusion: CD8+ T and Th1 cells, which secrete IFN-γ, might play important roles in the mucosal immunity of the stomach in children with H. pylori infection.

Effects of ß-glucan, probiotics, and synbiotics on obesity-associated colitis and hepatic manifestations in C57BL/6J mice.

PURPOSE: Probiotics and prebiotics are commonly used to improve the gut microbiota. Since prebiotics can support the growth of probiotics, co-administration of these is called synbiotics. It has been demonstrated that obesity-induced gut dysbiosis can worsen inflammatory bowel disease symptoms. This study evaluated how modulation of gut microbiota with Schizophyllum commune-derived ß-glucan (BG), probiotics (PRO), and synbiotics containing both BG and PRO (SYN) could improve the symptoms of obesity-associated colitis and hepatic manifestation. METHODS: Mice were fed a normal diet (ND), high-fat diet (HFD), and HFD with different additives (BG, PRO, and SYN) for 12 weeks, followed by 5 days of colitis induction. Mice were sacrificed before and after colitis induction. During the experiment, body weight, food and water consumption, and rectal bleeding were monitored. Proteins from the colon were subjected to western blotting, and serum biomarkers such as alanine transaminase, alkaline phosphatase, triglycerides, and total cholesterol were analyzed. Colon and liver samples were sectioned for histological analysis. The fecal microbiota was analyzed based on partial 16S rRNA gene sequences. RESULTS: Although BG and PRO secured intestinal tight junctions, these two treatments did not modulate inflammatory cell infiltration and inflammatory markers (i.e., IL-6 and TNF-α). In contrast, SYN demonstrated stronger and broader effects in reducing colonic inflammation. While BG treatment increased the abundance of indigenous Lactobacillus, PRO treatment decreased bacterial diversity by suppressing the growth of several species of bacteria. SYN treatment groups, however, supported the growth of both indigenous and supplemented bacteria while maintaining bacterial diversity. CONCLUSION: Obesity-associated colitis can be improved by modulating gut bacteria with ß-glucan and probiotics. The co-administration of both outperformed ß-glucan and probiotic treatment alone by fostering both indigenous and supplemented probiotic strains.

Cyr61 synthesis is induced by interleukin-6 and promotes migration and invasion of fibroblast-like synoviocytes in rheumatoid arthritis.

BACKGROUND: Interleukin-6 (IL-6) is involved in fibroblast-like synoviocyte (FLS) activation and promotes pannus formation and bone and cartilage destruction in rheumatoid arthritis (RA). Cysteine-rich 61 (Cyr61) protein regulates cell proliferation, migration, and differentiation. The aim of this study was to investigate the role of Cyr61 in RA-FLS migration and invasion after IL-6 stimulation. METHODS: Western blotting, immunohistochemistry, reverse transcription-polymerase chain reaction, and real time-polymerase chain reaction were used to examine protein and mRNA levels of Cyr61, matrix metalloproteinases (MMPs), and other signalling proteins. Knockdown of gene expression was performed with siRNA, and RNA sequencing was performed for differential gene analysis. Migration and invasion were assessed by wound healing and Boyden chamber assays. RESULTS: Cyr61 levels were elevated in FLSs from RA patients compared to those in osteoarthritis patients. Control and IL-6-treated FLSs showed differential gene expression. IL-6 stimulated protein synthesis of Cyr61, which was attenuated by the extracellular signal-related kinase 1/2 (ERK 1/2) inhibitor, PD98059, and knockdown of early growth response 3 (EGR3), but not of JUN. IL-6-induced Cyr61 protein synthesis increased expression of MMP2. Cyr61 promoted FLS migration and invasion in an autocrine manner. Knockdown of CYR61 and a neutralising antibody attenuated Cyr61 synthesis and IL-6-induced FLS migration. CONCLUSIONS: By modulating the ERK/EGR3 pathway, IL-6 stimulated Cyr61 production and in turn increased invasiveness of FLS. Our data suggest that Cyr61 might be a potential target to prevent the progression of joint damage in RA.

ß-Glucan-Based Wet Dressing for Cutaneous Wound Healing.

Objective: Recognized as pathogen-associated molecular patterns (PAMPs), ß-glucans, a naturally occurring heterogeneous group of polysaccharides, were investigated for their ability to accelerate wound healing in the form of high water-retaining hydrogel dressing. Approach: Full-thickness wounds on the dorsal side of mice created using a 5-mm biopsy punch were treated with ß-glucan-based hydrogel for 2 weeks. Standardized photographs of the wound site were taken at regular time intervals to calculate the percentage of wound closure. Tissues isolated from the wound area were subjected to histological examination and immunoblot analysis. Results: ß-Glucan-based hydrogel significantly accelerated the duration of wound healing and enhanced the development of skin appendages in the regenerated skin tissue. Increased expression of transforming growth factor-ß3 in the skin tissue isolated from the healed wound site indicated that skin regeneration rather than skin repair occurred, thereby minimizing cutaneous scarring. The expression level of cytokeratin 10 and cytokeratin 14 in the isolated skin tissue revealed that the wounds treated with hydrogel showed proper differentiation and proliferation of keratinocytes in the epidermal layer. Innovation: Immunomodulating ß-glucan (responsible for fighting infections at the wound site, and enhancing the migration and proliferation of keratinocytes and fibroblasts) in the form of a three-dimensional hydrogel membrane that retains a high water content (responsible for cooling and soothing effect around the wound site, thereby reducing pain) was prepared and analyzed for its effects on the cutaneous wound healing mechanism. Conclusion: ß-Glucan-based hydrogels are promising as wet wound dressings in the health care industry.

Mucosal Immunity Related to FOXP3+ Regulatory T Cells, Th17 Cells and Cytokines in Pediatric Inflammatory Bowel Disease.

BACKGROUND: We aimed to investigate mucosal immunity related to forkhead box P3 (FOXP3+) regulatory T (Treg) cells, T helper 17 (Th17) cells and cytokines in pediatric inflammatory bowel disease (IBD). METHODS: Mucosal tissues from terminal ileum and colon and serum samples were collected from twelve children with IBD and seven control children. Immunohistochemical staining was done using anti-human FOXP3 and anti-RORγt antibodies. Serum levels of cytokines were analyzed using a multiplex assay covering interleukin (IL)-1ß, IL-4, IL-6, IL-10, IL-17A/F, IL-21, IL-22, IL-23, IL-25, IL-31, IL-33, interferon (IFN)-γ, soluble CD40L, and tumor necrosis factor-α. RESULTS: FOXP3+ Treg cells in the lamina propria (LP) of terminal ileum of patients with Crohn's disease were significantly (P < 0.05) higher than those in the healthy controls. RORγt+ T cells of terminal ileum tended to be higher in Crohn's disease than those in the control. In the multiplex assay, serum concentrations (pg/mL) of IL-4 (9.6 ± 1.5 vs. 12.7 ± 3.0), IL-21 (14.9 ± 1.5 vs. 26.4 ± 9.1), IL-33 (14.3 ± 0.9 vs. 19.1 ± 5.3), and IFN-γ (15.2 ± 5.9 vs. 50.2 ± 42.4) were significantly lower in Crohn's disease than those in the control group. However, serum concentration of IL-6 (119.1 ± 79.6 vs. 52.9 ± 39.1) was higher in Crohn's disease than that in the control. Serum concentrations of IL-17A (64.2 ± 17.2 vs. 28.3 ± 10.0) and IL-22 (37.5 ± 8.8 vs. 27.2 ± 3.7) were significantly higher in ulcerative colitis than those in Crohn's disease. CONCLUSION: Mucosal immunity analysis showed increased FOXP3+ T reg cells in the LP with Crohn's disease while Th17 cell polarizing and signature cytokines were decreased in the serum samples of Crohn's disease but increased in ulcerative colitis.

A Novel Synthetic Material, BMM, Accelerates Wound Repair by Stimulating Re-Epithelialization and Fibroblast Activation.

Cutaneous wound repair is an intricate process whereby the skin reprograms itself after injury. In the mid-phase of wound repair, the proliferation, migration, and differentiation of cells are the major mechanisms to lead remodeling. We investigated the effect of BMM ((1E,2E)-1,2-bis((6-bromo-2H-chromen-3-yl)methylene)hydrazine), a novel synthetic material, on the migration and viability of keratinocytes or fibroblasts using the in vitro scratch woundhealing, electric cell-substrate imedance sensing (ECIS), invasion, and MTT assays. Cell migration-related factors were analyzed using western blot, and we found that treatment with BMM stimulated the EMT pathway and focal adhesion kinase (FAK)/Src signaling. Differentiation of HaCaT keratinocyte and fibroblast cells was also stimulated by BMM and specifically, NOX2/4 contributed to the activation of fibroblasts for wound healing. Furthermore, BMM treated HaCaT keratinocyte and fibroblast-co-cultured cells increased migration and differentiation. TGF-ß and Cyr61 were also secreted to a greater extent than in single cultured cells. In vivo experiments showed that treatment with BMM promotes wound closure by promoting re-epithelialization. In this study, we demonstrated that a novel synthetic material, BMM, is capable of promoting wound healing via the stimulation of re-epithelialization in the epidermis and the activation of fibroblasts in the dermis, in particular, via the acceleration of the interaction between the epidermis and dermis.

TMF and glycitin act synergistically on keratinocytes and fibroblasts to promote wound healing and anti-scarring activity.

Keratinocyte-fibroblast interactions are critical for skin repair after injury. During the proliferative phase of wound healing, proliferation, migration and differentiation of these cells are the major mechanisms leading to tissue remodeling. We have previously reported that glycitin, a major soy isoflavone, stimulates dermal fibroblast proliferation; and the phytochemical, 4',6,7-trimethoxyisoflavone (TMF), induces migration of HaCaT keratinocyte cells. We therefore investigated whether these compounds display synergistic effects on skin cells during wound healing in vitro and in vivo. Co-treatment with TMF and glycitin synergistically promotes the proliferation and migration of both keratinocytes and dermal fibroblasts, with a 1:1 ratio of these compounds showing the greatest efficacy in our co-culture system. This keratinocyte-fibroblast interaction occurred via the secretion of TGF-ß, and the induction of differentiation and proliferation was confirmed in both indirect and direct co-culture assays. In an excisional and burn wound animal model, mice treated with a 1:1 ratio of TMF and glycitin showed faster wound closure, regeneration and scar reduction than even the positive control drug. These data indicate that two isoflavones, TMF and glycitin, act synergistically to promote wound healing and anti-scarring and could potentially be developed together as a bioactive therapeutic for wound treatment.

Pazopanib for Non-small Cell Lung Cancer: The First Case Report in Korea.

Pazopanib is a potent multitargeted tyrosine kinase inhibitor that has been shown to have good efficacy in patients with renal cell carcinoma. A previous phase II trial demonstrated that short-term pazopanib administration was generally well tolerated and showed antitumor activity in patients with early-stage non-small cell lung cancer. Herein, we report on the case of a 66-year-old man with simultaneous metastatic squamous cell carcinoma of the lung and renal cell carcinoma who was treated with pazopanib. The patient showed an unexpected partial response and experienced a 10-month progression-free survival without significant toxicity. To the best of the authors' knowledge, this is the first report of pazopanib treatment in a non-small cell lung cancer patient in Korea. The results in this patient suggest that pazopanib may be a valid treatment option for advanced non-small cell lung cancer.

A case of exogenous corticosteroid-induced Kaposi's sarcoma that developed after a cure of endogenous hypercortisolism.

UNLABELLED: Case Kaposi's sarcoma (KS) is a malignant vascular tumor that occurs commonly in patients with acquired immunodeficiency syndrome. KS associated with Cushing's syndrome (CS) is unusual, especially in endogenous CS. Here, we report a case of KS associated with glucocorticoid-replacement therapy after surgical treatment for adrenal CS. A 70-year-old man presented with symptoms and signs of CS with a left adrenal mass. Adrenal CS was confirmed by biochemical studies. After left adrenalectomy, he took oral prednisolone (15 mg/day) to prevent adrenal insufficiency. Ten weeks later, numerous raised purple plaques on the lower extremities were newly detected. The biopsy findings were compatible with KS, but anti-HIV antibodies were negative. After withdrawal of glucocorticoid therapy, the skin lesions regressed completely. CONCLUSION: In this case, KS developed after the use of exogenous corticosteroid but not during endogenous hypercortisolism. This finding suggests that endogenous and exogenous corticosteroid play different roles in the development of KS.

Novel naphthochalcone derivative accelerate dermal wound healing through induction of epithelial-mesenchymal transition of keratinocyte.

BACKGROUND: Wound healing is an intricate process whereby the skin repairs itself after injury. The epithelial-mesenchymal transition (EMT) is associated with wound healing and tissue regeneration. Naphthochalcone derivatives have various pharmaceutical properties. We investigated the effect of a novel naphthochalcone derivative, 2-(5-(2,4,6-trimethoxyphenyl)-4,5-dihydro-1H-pyrazol-3-yl)naphthalen-1-ol (TDPN), on dermal wound healing in vivo and the migration of keratinocytes in vitro. RESULT: We investigated the effect of TDPN on signaling pathway and epithelial-mesenchymal transition through protein and transcriptional expression. The TDPN treatment accelerated dermal closure about 3 days and remodeling of dermis. We found that treatment with TDPN induced the migration of keratinocytes but not cytotoxicity. TDPN induced the phosphorylation of ERK and AKT. TDPN-treated cells showed loss of adherence protein and showed induction of the transcriptional factor Slug, mesenchymal marker, and fibronectin. Moreover, TDPN treatment induced the expression of matrix metalloproteinase-1 (MMP-1), which degrades specific components of the extracellular matrix, thereby providing new substrates that facilitate migration and invasion. MMP expression is considered to be one of the major attributes acquired by cells after EMT. CONCLUSION: We propose that a novel naphthochalcone derivative TDPN is capable of promoting keratinocyte migration via the induction of EMT resulting acceleration of wound closure and matrix remodeling.

Small cell carcinoma of the liver and biliary tract without jaundice.

An 80-year-old woman presenting with chest pain was found to have a large, lobulated soft tissue mass in the liver and nearby tissues on abdominal computed tomography (CT). The tumor had invaded the common hepatic artery and main portal vein. Jaundice developed 4 wk later, at which point, a pancreas and biliary CT scan revealed a large mass in the right lobe of the liver and a hilar duct obstruction, which was found to be a small cell carcinoma. Despite its rarity, liver and bile duct small cell carcinoma should be considered in the differential diagnosis of atypical chest pain without jaundice.

Inhibitory effects of Carpinus tschonoskii leaves extract on CpG-stimulated pro-inflammatory cytokine production in murine bone marrow-derived macrophages and dendritic cells.

This report describes the anti-inflammatory effects of MeOH extract from leaves of Carpinus tschonoskii (CE) on primary bone marrow-derived macrophage (BMDMs) and dendritic cells (BMDCs). Primary BMDMs and BMDCs were used for pro-inflammatory cytokine production and Western blot analysis. Human embryonic kidney cell line 293 T (HEK293 T) was used to access NF-κB activity. In all cases, CpG DNA was used to stimulate the cells. The CE (0-150 µg/ml) was treated to BMDMs, BMDCs, and HEK293T cells. CE pre-treatment in CpG-stimulated BMDMs and BMDCs showed a dose-dependent inhibitory effect on pro-inflammatory cytokine (e.g., IL-12 p40, IL-6, and TNF-α) production as compared to non-treated controls. The CE pre-treatment had no significant inhibition on mitogen-activated protein kinases (MAPKs) phosphorylation but strongly inhibited IκBα degradation. In NF-κB reporter gene assay, the CE pre-treatment inhibited NF-κB-dependent luciferase activity in a dose-dependent manner. Taken together, these data suggest that CE has significant inhibitory effect on pro-inflammatory cytokine production and warrant further studies concerning potentials of CE for medicinal uses.