AIM: To examine human β-defensin-3 (hBD-3) expression in inflamed gastric mucosal tissues or MKN45 gastric cancer cells with or without H pylori infection for better understanding the innate immune response to H pylo...AIM: To examine human β-defensin-3 (hBD-3) expression in inflamed gastric mucosal tissues or MKN45 gastric cancer cells with or without H pylori infection for better understanding the innate immune response to H pylori. METHODS: We used reverse transcription-polymerase chain reactions and immunohistochemistry to examine hBD-3 expression in inflamed gastric mucosal tissues or MKN45 gastric cancer cells with or without H pylori. Effects of hBD-3 against H pylori were also evaluated. RESULTS: The mean mRNA expression of hBD-3 in H pylori -positive specimens was significantly higher than that in H pylori-negative specimens (P = 0.0002, Mann-Whitney). In addition, unlike uninfected samples, 8 of 15 (53.33%) infected mucosal samples expressed hBD-3 protein. H pylori dose-dependently induced mRNA expression of hBD-3 in MKN45 cells, an effect inhibited by adding anti-toll-like receptor (TLR)-4 antibody. HBD-3 protein completely inhibited H pylori growth. CONCLUSION: Our results suggest that like hBD-2, hBD-3 may be involved in the pathophysiology of H pylori-induced gastritis.展开更多
To establish a cell line for stable expression of human beta-defensin 3 (hBD3). Methods Full length cDNA of hBD3 was isolated from previously constructed pGEM-hBD3 and then inserted into pcDNA3. The recombinant vector...To establish a cell line for stable expression of human beta-defensin 3 (hBD3). Methods Full length cDNA of hBD3 was isolated from previously constructed pGEM-hBD3 and then inserted into pcDNA3. The recombinant vector identified carrying hBD3 with right direction was introduced into COS-7 cells by Lipofe-ctamine. Cell clones survived in G418-rich medium and with stable expression of hBD3 in both mRNA and protein levels were identified by RT-PCR and Western blot respectively. Genomic integration of the hBD3 gene with the COS-7 cells was confirmed by Southern dot blot and primary analysis. The antimicrobial activity of the secreted hBD3 was also evaluated. Results COS-7 cells transfected with pcDNA3-hBD3 expressed hBD3 stably in mRNA and protein level. Southern dot blot analysis showed successful integration of the hBD3 gene into the genome of COS-7 cell and the hBD-3 protein secreted into the culture medium showed antimicrobial activity. Conclusion We successfully established a hBD3-expressing cell line.展开更多
Anti-microbial peptides are essential for the intestinal innate immunity that protects the intestinal epithelia from attacks by foreign pathogens. Human β-defensin (HBD) is one of the pivotal anti-microbial peptides ...Anti-microbial peptides are essential for the intestinal innate immunity that protects the intestinal epithelia from attacks by foreign pathogens. Human β-defensin (HBD) is one of the pivotal anti-microbial peptides that are expressed in the colonic epithelia. This study investigated the effect and the signaling mechanism of inducible β-defensin HBD2 by an essential amino acid, isoleucine (Ile) in colonic epithelial cells. Here we examined the expression level of HBD2 on induction of Ile in epithelial cells, and checked this pathway. HBD2 mRNA was induced by co-incubation with IL-1α and Ile in Caco2 cells, but not by Ile alone. An inhibitor of either ERK or Gi, a subunit of G-proteins, reduced the induction of HBD2 mRNA by Ile. The treatment with Ile also increased the intracellular calcium ion concentration, thus suggesting that the GPCR and ERK signaling pathway mediate the effects of Ile. These results indicate that an essential amino acid, Ile, enhances the expression of an inducible β-defensin, namely HBD2, by IL-1α through the activation of GPCRs and ERK signaling pathway. The administration of Ile may therefore represent a possible option to safely treat intestinal inflammation.展开更多
To investigate whether LL-37 and human beta defensin-2 (hBD-2) is related to the patients with psoriasis seldom having skin infections and explore the role of the two peptides and CCR6 (the receptor of hBD-2) in the p...To investigate whether LL-37 and human beta defensin-2 (hBD-2) is related to the patients with psoriasis seldom having skin infections and explore the role of the two peptides and CCR6 (the receptor of hBD-2) in the pathogenesis of psoriasis, the expression levels of mRNA of LL-37, hBD-2, and CCR6 in skin lesions of patients with psoriasis vulgaris were detected by using RT-PCR. The results showed that the mRNA expression levels of the two peptides and CCR6 in psoriatic lesions all increased compared with the normal skin (P<0.001). It was suggested that up-regulated expression of LL-37 and hBD-2 might be the main reason that result in the the skin of patients with psoriasis being seldom infected, and the two peptides and CCR6 might play crucial roles in the pathogenesis of psoriasis.展开更多
Amniochorion membranes were collected from 25 pregnant women at preterm labor, in the presence or not of Preterm Premature Rupture of Membranes (PPROM) and 27 pregnant women at term in the presence at labor, in order ...Amniochorion membranes were collected from 25 pregnant women at preterm labor, in the presence or not of Preterm Premature Rupture of Membranes (PPROM) and 27 pregnant women at term in the presence at labor, in order to quantify the expression and to evaluate the immunoreactivity of human beta defensins (HBD)1, HBD2, HBD3 and HBD4 in amniochorion membranes from pregnancies complicated by spontaneous prematurity. The HBDs were evaluated by immunohistochemistry, real time quantitative PCR and ELISA. Statistical analyses were performed using Chi-squared and Mann Whitney tests. There was no significant difference in HBDs expression between study and control groups: HBD1 (Md = 0.62 (0.0 - 105.0) vs Md = 0.80 (0.02 - 25.0);p = 0.85), HBD2 (Md = 0.17 (0.0 - 5.2) vs Md = 0.0 (0.0 - 43.2);p = 0.16), HBD3 (Md = 0.11 (0.0 - 140.5) vs Md = 0.06 (0.0 - 972.1);p = 0.91). Also, HBD1, HBD2 and HBD3 protein expression was not significant different between the groups: HBD1 (1.32 pg/mL (0.0 - 1.85) vs 1.08 pg/mL (0.04 - 2.22);p = 0.67), HBD2 (0.00 pg/mL (0.0 - 1.74) vs 0.02 pg/mL (0.0 - 1.24);p = 0.69), HBD3 (0.04 pg/mL (0.0 - 1.05) vs 0.09 pg/mL (0.0 - 1.05);p = 0.63). The immunoreactivity of HBD1, HBD2 and HBD3 was observed in amnion, chorion and decidua cells from preterm and term pregnancies. Amniochorion membranes are sources of HBD1, HBD2 and HBD3 and their expressions are similar in term and preterm pregnancies.展开更多
文摘AIM: To examine human β-defensin-3 (hBD-3) expression in inflamed gastric mucosal tissues or MKN45 gastric cancer cells with or without H pylori infection for better understanding the innate immune response to H pylori. METHODS: We used reverse transcription-polymerase chain reactions and immunohistochemistry to examine hBD-3 expression in inflamed gastric mucosal tissues or MKN45 gastric cancer cells with or without H pylori. Effects of hBD-3 against H pylori were also evaluated. RESULTS: The mean mRNA expression of hBD-3 in H pylori -positive specimens was significantly higher than that in H pylori-negative specimens (P = 0.0002, Mann-Whitney). In addition, unlike uninfected samples, 8 of 15 (53.33%) infected mucosal samples expressed hBD-3 protein. H pylori dose-dependently induced mRNA expression of hBD-3 in MKN45 cells, an effect inhibited by adding anti-toll-like receptor (TLR)-4 antibody. HBD-3 protein completely inhibited H pylori growth. CONCLUSION: Our results suggest that like hBD-2, hBD-3 may be involved in the pathophysiology of H pylori-induced gastritis.
文摘To establish a cell line for stable expression of human beta-defensin 3 (hBD3). Methods Full length cDNA of hBD3 was isolated from previously constructed pGEM-hBD3 and then inserted into pcDNA3. The recombinant vector identified carrying hBD3 with right direction was introduced into COS-7 cells by Lipofe-ctamine. Cell clones survived in G418-rich medium and with stable expression of hBD3 in both mRNA and protein levels were identified by RT-PCR and Western blot respectively. Genomic integration of the hBD3 gene with the COS-7 cells was confirmed by Southern dot blot and primary analysis. The antimicrobial activity of the secreted hBD3 was also evaluated. Results COS-7 cells transfected with pcDNA3-hBD3 expressed hBD3 stably in mRNA and protein level. Southern dot blot analysis showed successful integration of the hBD3 gene into the genome of COS-7 cell and the hBD-3 protein secreted into the culture medium showed antimicrobial activity. Conclusion We successfully established a hBD3-expressing cell line.
文摘Anti-microbial peptides are essential for the intestinal innate immunity that protects the intestinal epithelia from attacks by foreign pathogens. Human β-defensin (HBD) is one of the pivotal anti-microbial peptides that are expressed in the colonic epithelia. This study investigated the effect and the signaling mechanism of inducible β-defensin HBD2 by an essential amino acid, isoleucine (Ile) in colonic epithelial cells. Here we examined the expression level of HBD2 on induction of Ile in epithelial cells, and checked this pathway. HBD2 mRNA was induced by co-incubation with IL-1α and Ile in Caco2 cells, but not by Ile alone. An inhibitor of either ERK or Gi, a subunit of G-proteins, reduced the induction of HBD2 mRNA by Ile. The treatment with Ile also increased the intracellular calcium ion concentration, thus suggesting that the GPCR and ERK signaling pathway mediate the effects of Ile. These results indicate that an essential amino acid, Ile, enhances the expression of an inducible β-defensin, namely HBD2, by IL-1α through the activation of GPCRs and ERK signaling pathway. The administration of Ile may therefore represent a possible option to safely treat intestinal inflammation.
文摘To investigate whether LL-37 and human beta defensin-2 (hBD-2) is related to the patients with psoriasis seldom having skin infections and explore the role of the two peptides and CCR6 (the receptor of hBD-2) in the pathogenesis of psoriasis, the expression levels of mRNA of LL-37, hBD-2, and CCR6 in skin lesions of patients with psoriasis vulgaris were detected by using RT-PCR. The results showed that the mRNA expression levels of the two peptides and CCR6 in psoriatic lesions all increased compared with the normal skin (P<0.001). It was suggested that up-regulated expression of LL-37 and hBD-2 might be the main reason that result in the the skin of patients with psoriasis being seldom infected, and the two peptides and CCR6 might play crucial roles in the pathogenesis of psoriasis.
文摘Amniochorion membranes were collected from 25 pregnant women at preterm labor, in the presence or not of Preterm Premature Rupture of Membranes (PPROM) and 27 pregnant women at term in the presence at labor, in order to quantify the expression and to evaluate the immunoreactivity of human beta defensins (HBD)1, HBD2, HBD3 and HBD4 in amniochorion membranes from pregnancies complicated by spontaneous prematurity. The HBDs were evaluated by immunohistochemistry, real time quantitative PCR and ELISA. Statistical analyses were performed using Chi-squared and Mann Whitney tests. There was no significant difference in HBDs expression between study and control groups: HBD1 (Md = 0.62 (0.0 - 105.0) vs Md = 0.80 (0.02 - 25.0);p = 0.85), HBD2 (Md = 0.17 (0.0 - 5.2) vs Md = 0.0 (0.0 - 43.2);p = 0.16), HBD3 (Md = 0.11 (0.0 - 140.5) vs Md = 0.06 (0.0 - 972.1);p = 0.91). Also, HBD1, HBD2 and HBD3 protein expression was not significant different between the groups: HBD1 (1.32 pg/mL (0.0 - 1.85) vs 1.08 pg/mL (0.04 - 2.22);p = 0.67), HBD2 (0.00 pg/mL (0.0 - 1.74) vs 0.02 pg/mL (0.0 - 1.24);p = 0.69), HBD3 (0.04 pg/mL (0.0 - 1.05) vs 0.09 pg/mL (0.0 - 1.05);p = 0.63). The immunoreactivity of HBD1, HBD2 and HBD3 was observed in amnion, chorion and decidua cells from preterm and term pregnancies. Amniochorion membranes are sources of HBD1, HBD2 and HBD3 and their expressions are similar in term and preterm pregnancies.
