당뇨 백서 사구체 및 고농도 포도당으로 자극한 족세포에서 ZO-1의 변화와 안지오텐신 수용체 차단제의 영향

김범석 ( Kim Beom Seog ),허종호 ( Heo Jong Ho ),박형천 ( Park Hyeong Cheon ),강신욱 ( Kang Sin Ug ),최규헌 ( Choe Gyu Heon ),하성규 ( Ha Seong Gyu ),이호영 ( Lee Ho Yeong ),한대석 ( Han Dae Seog ) 대한신장학회 2003 Kidney Research and Clinical Practice Vol.22 No.6

목 적 : 당뇨병성 신증은 임상적으로 단백뇨가 특징적인 소견으로, 최근의 연구에 의하면 당뇨병성 신증을 포함한 대부분의 사구체 질환에 의한 단백뇨는 사구체 여과 장벽의 이상에 의해 발생하는 것으로 알려져 있다. 본 연구에서는 고농노 포도당으로 자극한 배양 족세포와 streptozotocin으로 당뇨가 유발된 백서의 사구체를 이용하여 slit diaphragm과 연관이 있는 세포질 내 단백 중 하나인 zona occludens-1 (ZO-1)의 변화 및 안지오텐신 제 1형 수용체 차단제 (angiotensin Ⅱ type 1 receptor blocker, ARB)의 영향에 대해 알아보고자 하였다. 방 법 : 불멸화 마우스 족세포 (immortalized mouse podocytes)를 정상 포도당 (5.6 mM, NG), Ng^(+) 만니콜 24.4 mM (NG+M), NG+ARB (L-158,809 10^(6)M), 고농도 포도당 (30 mM, HG) 및 HG+ARB로 7일간 자극한 후 ZO-1 mRNA와 단백의 변화를 RT-PCR과 western blot을 이용하여 관찰하였다. 또한, 54마리의 Sprague-Dawley 백서를 대상으로 대조군 (18마리), 당뇨군 (18마리) 및 ARB 투여 당뇨균 (18마리)으로 나눈 후 실험 1,2,3개월에 사구체를 분리하여 RT-PCR과 Western blot을 시행하였으며, 조직에서의 ZO-1 발현은 면역조지화학 염색법을 이용하여 관찰하였다. 결 과 : 7일간 고농도 포도당 배양액으로 배양한 족세포 내의 ZO-1 mRNA와 단백 발현은 정상 포도당으로 배양한 족세포에 비하여 각각 283±22%와 220±12% 증가되었으며 (p<0.01), ARB를 투여한 고농도 포도당 배양액으로 배양한 족세포에서는 고농도 포도당군에서 관찰된 ZO-1 mRNA와 단백 발현의 증가가 각각 73±9%와 64±7% 억제되었다 (p<0.05). 24시간 요단백 배설량은 당뇨균에서 1개월 후 78.7±7.2 ㎎ 72.8±7.2 ㎎, 그리고 3개월 후 90.6±11.3㎎으로 대조군의 1개월 후 23.0±1.5㎎, 2개월 후 23.9±1.2㎎, 및 3개월 후 19.8±3.2 ㎎에 비해 의의있게 증가되었으며 (p<0.01), 이러한 증가는 ARB의 투영에 의해 1개월 후 56.3±6.0 ㎎ 2개월후 52.5±3.6 ㎎ 및 3개월 후 51.6±56.6 ㎎로 의의있게 억제되었다 (p<0.05). 당뇨군에서 사구체 내 ZO-1 mRNA 및 단백 발현은 대조군에 비해 모든 기간에서 의의있게 증가되었으며, 특히 3개월 후에는 각각 234±19%와 216 ±20% 증가되었고 (p<0.01), 이러한 증가는 ARB의 투여로 각각 57±9%와 71±14% 감소하였다 (p<0.05). 면역조직화학 검사상 당뇨군의 사구체에서 대조군에 비해 ZO-1 단백의 발현이 의의있게 증가되었으며, ARB 투여군에서는 당뇨군에 비해 의의있게 억제되었다. 이러한 ZO-1 발현의 변화는 주로 족세포에서 관찰되었다. 결 론 : 이상의 생체내외 실험 결과로부터, 족세포 내의 ZO-1 발현의 변화가 당뇨병성 신증의 초기부터 나타나며, 이러한 변화는 단백뇨의 발생과 연관성이 있을 것으로 생각된다. Background : Diabetic nephropathy (DN) is clinically characterized by persistent proteinuria. The underlying pathologic changes responsible for the nephropathy are the loss of size selective and/or charge seletive properties of the glomerular filtration harrier. Size selectivity is maintained primarily by the slit diaphragm and ZO-1 is one of the basic components of it. However, the precise role of the ZO-1 in the pathogenesis of the glomerular diseases is not fully understood. We investigated the changes of ZO-1 expression in diabetic glomeruli in vivo. We also evaluated the effect of angiotensin Ⅱ type 1 receptor blocker (ARB) on the ZO-1 changes induced by diabetes or high glucose. Methods : To determine the effect of ARB on podocytes ZO-1 protein and mRNA expression, immortalized mouse podocytes were incubated with RPMI medium containing mormal glucose (NG, 5.6mM) or high glucose (HG, 30 mM) with or without ARB (10?? M, L-158,809). For animal studies, rats were injected with diluent (Control, C, n=18) or streptozotocin. The latter were left untreated (DM, n=18) or treated with 1 mg/㎏/day ARB (DM+ARB, n=18). Six rats from each group were sacrificed monthly, and Western blot and RT PCR were performed for ZO-1 with sieved glomeruli. Renal sections were stained for ZO-1 by immunohistochemistry. Results : The ZO-1 mRNA and protein expressions in podocytes exposed to HG conditions were significantly higher than those in podocytes exposed to NG media (p<0.05). ARB treatment inhibited the HG induced increase in ZO-1 mRNA and protein expression by 73% and 64%, respectively (p<0.05). Compared to the C rats (19.8+3.2mg/day), 24 hour urinary protein excretion at 3 month was significantly higher in the DM rats (90.6±11.3 mg/day, p<0.05), and ARB treatment partly reversed the in crease in proteinuria in DM rats (51.6±6.6 mg/day, p<0.05). Glomerular ZO-1 mRNA and protein expressions were also significantly increased in DM than corresponding C at all duration (p<0.05). ARB treatment for 3 months in DM rats inhibited the increase in ZO-1 mRNA and protein expression by 57.5% and 70.6%, respectively (p<0.05). ARB treatment for 3 months significantly ameliorated increased glomerular ZO-1 expression in DM rats as assessed by immunohistochemistry. Conclusion : In conclusion, ZO-1 mRNA and protein expressions were increased in podocytes exposed to HG and in DM glomeruli, and this increment in ZO-1 expression was ameliorated with ARB. Taken together, these data suggest that change of ZO-1 expression in podocytes is implicated in the early changes of diabetic nephropathy and may contribute to the development of proteinuria. (Korean J Nephrol 2003;22(6):632-644)

당과 후기당화합물에 의한 사구체 상피세포 ZO-1 발현의 변화

이진석,이해수,윤옥자,하태선,Lee Jin-Seok,Lee Hae- Soo,Yoon Ok-Ja,Ha Tae-Sun 대한소아신장학회 2004 Childhood kidney diseases Vol.8 No.2

목적: 단백뇨 질환에서 볼 수 있는 사구체 상피세포(glomerular epithelial cells, GEpC) 족돌기(foot process)의 병리학적 변화에 있어서 GEpC사이의 세극막(slit diaphragm)과 세포골격을 연결하는 ZO-1 단백의 당뇨조건에 따른 변화를 알아보고자 하였다. 방법: 백서 GEpC을 배양하고 고농도의 당과 후기당화합물(advanced glycosylation enduroducts, AGE)를 적용하여 당뇨병 환경에 가까운 조건을 설정한 후, ZO-1 단백양은 Western 분석으로, 분포 변화는 공초점 현미경으로, 유전자 표현의 변화는 RT-PCR로 관찰하였다. 실험군은 당의 농도를 5 또는 30 mM로, AGE와 BSA를 첨가하고 osmotic control로서 당 5 mM에 mannitol 25 M을 섞은 것을 조합하여 A5, A30, B5, B30, Aosm로 하였다. 결과: 공초점 현미경 상 ZO-1은 정상적인 환경인 B5에서 B30, A5, 가장 병적인 A30 환경으로 진행할수록 세포질의 바깥에서 안쪽으로 이동하는 양상을 보였다. ZO-1 단백양은 B5 결과를 대조군으로 비교하여 당을 첨가한 B30에서 11.1%, AGE를 추가한 조건인 A5에서 2.3% 감소하였나 통계적 유의성은 없었고, 당과 AGE가 동시에 첨가된 A30에서는 19.0%의 유의한 감소를 보였다. mRNA의 발현도 A30에서만 12.0%의 의의 있는 감소를 보였다. 이러한 단백질과 mRNA의 감소 소견은 osmotic control (Aosm)에서는 관찰할 수 없었다. 결론: 고농도의 당과 AGE에 의한 GEpC의 ZO-1의 분포 변화와 유전자 수준에서의 억제로 단백의 생성 감소를 초래함으로써, 장기간 당뇨 환경체서 족돌기의 형태학적 변화를 설명할 수 있으며, 추후 이의 변화 기전에 대한 연구가 필요할 것으로 사료된다. Purpose: Regardless of the underlying diseases, the proteinuric condition demonstrates ultrastructural changes in podocytes with retraction and effacement of the highly specialized interdigitating foot processes. We examined the molecular basis for this alteration of the podocyte phenotypes, including quantitative and distributional changes of ZO-1 protein as a candidate contributing to the pathogenic changes in the barrier to protein filtration. Methods: To investigate whether high glucose and advanced glycosylation endproduct(AGE) induce podocyte cytoskeletal changes, we cultured rat GEpC under 1) normal glucose(5 mM=control) or 2) high glucose(30 mM) or 3) AGE-added or 4) high glucose plus AGE-added conditions. The distribution of ZO-1 was observed by confocal microscope and the change of ZO-1 expression was measured by Western blotting and RT-PCR. Results: By confocal microscopy, we observed that ZO-1 moves from peripheral cytoplasm to inner actin filaments complexes in both AGE-added and high glucose condition. In Western blotting, high glucose or AGE-added condition decreased the ZO-1 protein expression by 11.1%(P>0.05) and 2.3%(P>0.05), respectively compared to the normal glucose condition. High glucose plus AGE-added condition further decreased ZO-1 protein expression to statistically significant level(12%, P<0.05). No significant change was seen in the osmotic control. In RT-PCR, high glucose plus AGE-added condition significantly decreased the expression of ZO-1 mRNA by 12% compared to normal glucose condition. Conclusion: We suggest that both high glucose and AGE-added condition induce the cytoplasmic translocation and suppresses the production of ZO-1 at transcriptional level and these changes may explain the functional changes of podocytes in diabetic conditions.

유방암에서 Claudin-1과 ZO-1 단백 발현 감소와 임상-병리학적 인자와의 연관성

김민준(Min Joon Kim),차성재(Seong Jae Cha),박성일(Sung Il Park),박성준(Sung Jun Park),장인택(In Taik Chang),김범규(Beom Gyu Kim),최유신(Yoo Shin Choi),이태진(Tae Jin Lee) 대한외과학회 2007 Annals of Surgical Treatment and Research(ASRT) Vol.72 No.6

Purpose: Tight junction proteins are either over-expressed or suppressed in some cancers. These suppressions of claudin-1 and ZO-1 protein are known to have a significant relationship with the progression of breast cancer. The authors reviewed 42 cases of breast cancer and the staining status of claudin-1 and ZO-1 in order to evaluate Claudin-1 and ZO-1 as clinicopathologic risk factors. Methods: Immunohistological staining for Claudin-1 and ZO-1 was performed in 42 post-operative pathologically diagnosed infiltrating duct carcinoma specimens. The rate of expression was compared with the clinical record, the pathological diagnosis, the estrogen receptor and progesteron receptor status and the c-erbB2 gene to evaluate the protein expression-breast cancer progression relationship and to investigate the expressions of Claudin-1 and ZO-1 as a prognostic factors in breast cancer. Results: The claudin-1 and ZO-1 expressions were both decreased in all the post-operative specimens. The claudin-1 expressions were significantly decreased 100%, 82.4% and 66.7% as the histologic grade increased. The ZO-1 expressions were shown in 44.8% of the lymph node metastasis negative group and in 7.7% of the lymph node positive group. The expression of ZO-1 decreased by 53.3%, 28.6% and 0%, with statistical significance, as the stage increased. Conclusion: The claudin-1 expressions were decreased in the poorly differentiated group, i.e., a high histologic grade, and the ZO-1 expressions were decreased in the lymph node positive group and in the high stage cancer. This shows the possibility of Claudin-1 and ZO-1 as factors for tumorigenesis and progression and as prognostic factors in breast cancer.

Lactobacillus casei LC01 Regulates Intestinal Epithelial Permeability through miR-144 Targeting of OCLN and ZO1

( Qiuke Hou ),( Yongquan Huang ),( Yan Wang ),( Liu Liao ),( Zhaoyang Zhu ),( Wenjie Zhang ),( Yongshang Liu ),( Peiwu Li ),( Xinlin Chen ),( Fengbin Liu ) 한국미생물 · 생명공학회 2020 Journal of microbiology and biotechnology Vol.30 No.10

Our previous report determined that miR-144 is a key regulator of intestinal epithelial permeability in irritable bowel syndrome with diarrhea (IBS-D) rats. Recent evidence has shown that lactobacilli play an important role in the relief of IBS-D symptoms. However, few studies have addressed the mechanisms by which microRNAs and lactobacilli exert their beneficial effects on intestinal epithelial permeability. Hence, to elucidate whether miRNAs and lactobacilli play roles in intestinal epithelial barrier regulation, we compared miRNA expression levels in intestinal epithelial cells (IECs) under Lactobacillus casei (L. casei LC01) treatment. IECs and L. casei LC01 were co-cultured and then subjected to microRNA microarray assay. qRT-PCR, western blot and ELISA were used to detect the expression of occludin (OCLN) and zonula occludens 1 (ZO1/TJP1). The interaction between miRNAs and L. casei LC01 acting in IECs was investigated through transfection of RNA oligoribonucleotides and pcDNA 3.1 plasmid. The results are as follows: 1) L. casei LC01 decreased the expression of miR-144 and FD4 and promoted OCLN and ZO1 expression in IECs; 2) L. casei LC01 enhanced the barrier function of IECs via downregulation of miR-144 and upregulation of OCLN and ZO1; 3) Under L. casei LC01 treatment, OCLN and ZO1 overexpression could partially eliminate the promoting effect of miR-144 on intestinal permeability in IECs. Our results demonstrate that L. casei LC01 regulates intestinal permeability of IECs through miR-144 targeting of OCLN and ZO1. L. casei LC01 can be a possible therapeutic target for managing dysfunction of the intestinal epithelial barrier.

Extracellular domain of V-set and immunoglobulin domain containing 1 (VSIG1) interacts with Sertoli cell membrane protein, while its PDZ-binding motif forms a complex with ZO-1

Ekyune Kim,Youngjeon Lee,Kyu-Tae Chang 한국발생생물학회 2010 한국발생생물학회 학술발표대회 Vol.29 No.-

Mammalian spermatogenesis takes place in the seminiferousepithelium, which is composed of Sertoli cells and germ cells. The interaction between spermatogenic and Sertoli cells as well as elongated spermatids and Sertoli cells is tightly regulated by junctional adhesion molecules (JAMs). JAMs, which are cell adhesion molecules, are known to play roles in various biological processes such as fertilization, neurogenesis, cancer progression, and spermatogenesis. Members of the JAM family have a unique structure: they contain an N-terminal signal peptide domain, immunoglobulin (Ig)-like domains, transmembrane and cytoplasmic tail domains, each of which has distinct functions. The extracellular Ig-like domains interact in a homophilic or heterophilic manner, whereas cytoplasmic tail domain mediates the tight junction assembly. Although members of the JAM family are exclusively present in or restricted to the testis, their precise roles in spermatogenesis and fertilization have not yet been completely explored. The functional roles of Nectin-2, Nectin-3, JAM-C, cell adhesion molecule1 (CADM1), coxsackie and adenovirus receptor (CAR) have been evaluated by analysis of null mutant mice. Unfortunately, CAR-deficient mice had an embryonic lethal phenotype; this demonstrates the importance of CAR in development, but its physiological role in spermatogenesis is not known. The loss of CADM1, Nectin-3 and JAM-C resulted in male infertility caused by loss of adhesion between germ and Sertoli cells. A variety of JAMs participate in the interaction between germ and Sertoli cells. Recently, human VSIG1 has been characterized, which was originally known as A34, as a new member of the JAM family; VSIG1 is composed of two extracellular Ig-like domains, a transmembrane domain, and a cytoplasmic domain. However, this molecule has not been functionally characterized, so this was one of the aims of our present study. RT-PCR and immunoblot analyses were used to study VSIG1 expression, VSIG1 was specifically expressed in testicular germ cells but not in sperm. Pull-down assay with glutathione S-transferase (GST) or His-fused first Ig and second Ig domains of VSIG1 and SDS-PAGE under mild non-reducing conditions demonstrated that VSIG1 functions as an in vitro homophilic adhesion molecule. Furthermore, cells expressing a deletion of the C-terminus of VSIG1 failed to interact with ZO-1, the central structural protein of the tight junction. These findings suggest mouse VSIG1 interacts with an unknown molecule in Sertoli cells via its extracellular domain, while its cytoplasmic domain is needed for binding to ZO-1. Thus, we suggest mouse VSIG1 may play an important role in spermatogenesis rather than fertilization by forming heterophilic complex with a molecule similar to JAM family.

Nuclear Factor I-C 결손 생쥐에서 상아모세포의 형태학적 특징

고승백,이창섭,이난영,이상호,김흥중,박주철 大韓小兒齒科學會 2006 大韓小兒齒科學會誌 Vol.33 No.2

NFI-C K/O 생쥐에서는 상아모세포의 분화과정에 이상이 초래되어 상아질 형성에 이상이 생기고 치근 형성이 불완전하게 이루어지는 것으로 알려져 있으나 이에 대한 명확한 기전은 잘 알려져 있지 않다. 상아모세포가 분화하여 정상적으로 상아질을 형성하기 위해서 핵과 세포질이 극성을 띠고 잘 조직화되어야 하며, 이 과정에서 다양한 세포사이 결합장치들이 중요한 역할을 하는 것으로 알려져 있다. 본 연구에서는 NFI-C K/O 생쥐에서 상아질 형성에 이상이 생기는 것이 상아포세포의 형태학적 변화와 세포사이 결합장치들이 기능을 하지 못한 결과에서 기인한 것인지 알아보기 위하여, NFI-C K/O 생쥐에서 발생한 비정상적인 상아모세포들을 광학 및 투과 전자현미경을 이용하여 형태학적으로 관찰하고, Zo-1과 occludin의 발현을 면역조직화학적으로 관찰하여 세포사이 결합장치들의 분포를 확인하여 다음과 같은 결과를 얻었다. 1. 광학현미경 소견에서 NFI-C K/O 생쥐의 전치부 상아모세포는 세포 극성이 상실되고, 여러 층으로 배열되어 있었으며 상아질에 많은 세포들이 함입된 것과 같은 비정상적인 상아질의 소견을 나타냈다. 반면에 NFI-C K/O 생쥐의 구치부 상아모세포는 치관부에서는 잘 조직화된 소견을 보였으나 치근 형성 부위에서는 세포 배열이 불규칙해지고 세포 극성이 상실되었다. 2. 투과 전자현미경 소견에서 NFI-C K/O 생쥐 전치부의 비정상적인 상아모세포는 둥근 형태로 세포 사이 간격이 넓으며 폐쇄연접과 같은 세포사이 결합장치들이 전혀 관찰되지 않았다. 3. Zo-1의 면역조직화학적 염색에서 NFI-C K/O 생쥐의 전치부 법랑모세포의 근위부와 원위부에서 ZO-1이 강하게 발현되었으나 비정상적인 상아모세포에서는 ZO-1의 발현을 관찰할 수 없었다. 4. Occludin의 면역조직화학적 염색에서 정상 생쥐의 전치부 상아모세포에서는 occludin의 발현이 관찰되었으나 NFI-C K/O 생쥐의 비정상적인 상아모세포에서는 occludin의 발현이 관찰되지 않았다. 이상의 결과를 종합하여 볼 때 NFI-C의 결손은 상아모세포의 분화 이상을 초래하고 비정상적으로 상아질을 형성하는 과정에 세포사이 결합장치의 상실과 같은 형태학적인 변화의 중요한 요소로 작용하는 것으로 생각된다. NFO-C null mice demonstrated aberrant odntoblast differentiation and thus abnormal dentin formation while other tissues/organs in the body, including ameloblasts, appear to be unaffected and normal. However, little is known about the mechanism of NFI-C function in odontoblast differentiation and dentin formation. Odontoblasts are tall, highly polarized cells that are responsible for formation and maintenance of the predentin and dentin. An indication of their polarity is the acquisition of specialized intercellular junctions. As predontoblasts differentiate into odontoblasts, they are joined and attached at the apical end by well developed terminal webs of cytoskeletal actins, and associated tight as well as adherent njunctions. In this study, in order to investigate if disruption of the NFI-C gene interferes with formation of a specific or other structural proteins of the intercellular junctions, we examined morphological charcteristic of the aberrant odontoblast in NFI-C null mice using light and electron microscope. In addition, we determined the expression of major structural proteins of intercellular junctions, ZO-1 and occludin, during the differentiation of odontoblasts using immunohitochemistry. The results were as follows: 1. In light microscopy, abnormal odontoblasts of incisors of the NFI-C null mice were round in shape, lost their polarity, and trapped in osteodentin-like mineralized tissue. Mutant molars have relatively normal crowns, but short and abnormal differentiating adontoblasts in root formation area. 2. Electron microscopy of abnormal odontoblasts revealed the dissociation of the round osteoblast-like cells, the loss of their cellular polarity, and the absence of an intercellular junctional complex known as the tight junctions. 3. A mutant incisor showed labeling for ZO-1 at the proximal and distal ends of secreting ameloblasts, while staining for ZO-1 was not observed in the abnormal odontoblasts. 4. A normal incisor showed immunoreactivity for occludin in the differentiating odontoblasts. However, staining for occludin was not observed in the abnormal odntoblasts of mutant incisor. These results suggest that NFI-C gene causes dissociation of odontoblast and thus abberant odontoblast differentiation and abnormal dentin formation by interfering with the formation of intercellular junctions.

Ginseng Total Saponin Improves Podocyte Hyperpermeability Induced by High Glucose and Advanced Glycosylation Endproducts

Ha, Tae-Sun,Choi, Ji-Young,Park, Hye-Young,Lee, Jin-Seok The Korean Academy of Medical Sciences 2011 JOURNAL OF KOREAN MEDICAL SCIENCE Vol.26 No.10

<P>Early diabetic nephropathy is characterized by glomerular hyperpermeability as a result of impaired glomerular filtration structure caused by hyperglycemia, glycated proteins or irreversible advanced glycosylation endproducts (AGE). To investigate the effect of ginseng total saponin (GTS) on the pathologic changes of podocyte ZO (zonula occludens)-1 protein and podocyte permeability induced by diabetic conditions, we cultured mouse podocytes under: 1) normal glucose (5 mM, = control); 2) high glucose (HG, 30 mM); 3) AGE-added; or 4) HG plus AGE-added conditions and treated with GTS. HG and AGE increased the dextran filtration of monolayered podocytes at early stage (2-8 hr) in permeability assay. In confocal imaging, ZO-1 colocalized with actin filaments and β-catenin at cell contact areas, forming intercellular filtration gaps. However, these diabetic conditions suppressed ZO-1 immunostainings and disrupted the linearity of ZO-1. In Western blotting, diabetic conditions also decreased cellular ZO-1 protein levels at 6 hr and 24 hr. GTS improved such quantitative and qualitative changes. These findings imply that HG and AGE have an influence on the redistribution and amount of ZO-1 protein of podocytes thereby causing hyperpermeability at early stage, which can be reversed by GTS.</P>

Ginseng Total Saponin Improves Podocyte Hyperpermeability Induced by High Glucose and Advanced Glycosylation Endproducts

하태선,최지영,박혜영,이진석 대한의학회 2011 Journal of Korean medical science Vol.26 No.10

Early diabetic nephropathy is characterized by glomerular hyperpermeability as a result of impaired glomerular filtration structure caused by hyperglycemia, glycated proteins or irreversible advanced glycosylation endproducts (AGE). To investigate the effect of ginseng total saponin (GTS) on the pathologic changes of podocyte ZO (zonula occludens)-1 protein and podocyte permeability induced by diabetic conditions, we cultured mouse podocytes under: 1) normal glucose (5 mM, = control); 2) high glucose (HG, 30 mM); 3) AGE-added;or 4) HG plus AGE-added conditions and treated with GTS. HG and AGE increased the dextran filtration of monolayered podocytes at early stage (2-8 hr) in permeability assay. In confocal imaging, ZO-1 colocalized with actin filaments and β-catenin at cell contact areas, forming intercellular filtration gaps. However, these diabetic conditions suppressed ZO-1 immunostainings and disrupted the linearity of ZO-1. In Western blotting, diabetic conditions also decreased cellular ZO-1 protein levels at 6 hr and 24 hr. GTS improved such quantitative and qualitative changes. These findings imply that HG and AGE have an influence on the redistribution and amount of ZO-1 protein of podocytes thereby causing hyperpermeability at early stage, which can be reversed by GTS.

Curcumin protects against the intestinal ischemia-reperfusion injury: involvement of the tight junction protein ZO-1 and TNF-α related mechanism

Shuying Tian,Ruixue Guo,Sichen Wei,Yu Kong,Xinliang Wei,Weiwei Wang,Xiaomeng Shi,Hongyu Jiang 대한생리학회-대한약리학회 2016 The Korean Journal of Physiology & Pharmacology Vol.20 No.2

Present study aimed to investigate the effect of curcumin-pretreatment on intestinal I/R injury and on intestinal mucosa barrier. Thirty Wistar rats were randomly divided into: sham, I/R, and curcumin groups (n=10). Animals in curcumin group were pretreated with curcumin by gastric gavage (200 mg/kg) for 2 days before I/R. Small intestine tissues were prepared for Haematoxylin & Eosin (H&E) staining. Serum diamine oxidase (DAO) and tumor necrosis factor (TNF)-α levels were measured. Expression of intestinal TNF-α and tight junction protein (ZO-1) proteins was detected by Western blot and/or immunohistochemistry. Serum DAO level and serum and intestinal TNF-α leves were significantly increased after I/R, and the values were markedly reduced by curcumin pretreatment although still higher than that of sham group (p<0.05 or p<0.001). H&E staining showed the significant injury to intestinal mucosa following I/R, and curcumin pretreatment significantly improved the histological structure of intestinal mucosa. I/R insult also induced significantly down-regulated expression of ZO-1, and the effect was dramatically attenuated by curcumin-pretreatment. Curcumin may protect the intestine from I/R injury through restoration of the epithelial structure, promotion of the recovery of intestinal permeability, as well as enhancement of ZO-1 protein expression, and this effect may be partly attributed to the TNF-α related pathway.

Curcumin protects against the intestinal ischemia-reperfusion injury: involvement of the tight junction protein ZO-1 and TNF-α related mechanism

Tian, Shuying,Guo, Ruixue,Wei, Sichen,Kong, Yu,Wei, Xinliang,Wang, Weiwei,Shi, Xiaomeng,Jiang, Hongyu The Korean Society of Pharmacology 2016 The Korean Journal of Physiology & Pharmacology Vol.20 No.2

Present study aimed to investigate the effect of curcumin-pretreatment on intestinal I/R injury and on intestinal mucosa barrier. Thirty Wistar rats were randomly divided into: sham, I/R, and curcumin groups (n=10). Animals in curcumin group were pretreated with curcumin by gastric gavage (200 mg/kg) for 2 days before I/R. Small intestine tissues were prepared for Haematoxylin & Eosin (H&E) staining. Serum diamine oxidase (DAO) and tumor necrosis factor (TNF)-${\alpha}$ levels were measured. Expression of intestinal TNF-${\alpha}$ and tight junction protein (ZO-1) proteins was detected by Western blot and/or immunohistochemistry. Serum DAO level and serum and intestinal TNF-${\alpha}$ leves were significantly increased after I/R, and the values were markedly reduced by curcumin pretreatment although still higher than that of sham group (p<0.05 or p<0.001). H&E staining showed the significant injury to intestinal mucosa following I/R, and curcumin pretreatment significantly improved the histological structure of intestinal mucosa. I/R insult also induced significantly down-regulated expression of ZO-1, and the effect was dramatically attenuated by curcumin-pretreatment. Curcumin may protect the intestine from I/R injury through restoration of the epithelial structure, promotion of the recovery of intestinal permeability, as well as enhancement of ZO-1 protein expression, and this effect may be partly attributed to the TNF-${\alpha}$ related pathway.