The Preliminarily Basic Study On Mechanism Of Kidney Stone Under Metabolic Syndrome Environment

Part1：AParacrine Loop involving renal tubular cells,Adipocytes andMacropages Aggravates Inflammatory Changes to promote kidney stoneformation in metabolic syndromeBackground：Urolithiasis is a common urological disorder with a lifetime risk of10–12%among thepopulations of industrialized countries. In particular, the prevalence of kidney stones hasincreased, worldwide, over the past few decades. Dietary and lifestyle factors play anincreasing role in the risk of stone disease, with diets high in protein and fat raising the riskof stone formation. Recent studies have indicated that nephrolithiasis is linked to otherchronic diseases, such as diabetes mellitus, obesity and metabolic syndrome (MetS).MetS involves a constellation of manifestations, including visceral fat obesity, impairedglucose metabolism, atherogenic dyslipidemia, and hypertension. There is compellingevidence that obesity is a crucial etiologic factor in the development of MetS, with obesityalso being considered a metabolic disease and a chronic, low-grade, inflammatory disease.Recent studies have revealed that adipose tissue in MetS is characterized by the infiltrationof macrophages, which are responsible for the production of inflammatory cytokines.Despite a large amount of epidemiologic evidence supporting the association betweenMetS and kidney stone formation, the mechanism linking MetS with the formation ofkidney stones is largely unknown. Our previous research in mice showed that MetSaggravates the formation of calcium oxalate kidney stones by enhancing inflammation. Tofurther elucidate how MetS promotes susceptibility to urolithiasis before and during kidneystone formation, we simulated MetS by coculturing renal tubular epithelial cells with adipocytes and/or macrophages. We hypothesized that adipocytes and macrophagesincreased their cellular interactions to accelerate stone formation.Objective:The aim of this study is to establish an in vitro experimental system composed of renaltubular cells with adipocytes and macrophages to simulate metabolic syndrome, and thenexamine the molecular mechanism whereby these cells communicate.Methods:Mouse renal tubular cells (M-1s) is cocultured with adipocytes (3T3-L1s) or/andmacrophages (RAW264s). The calcium oxalate monohydrate (COM) crystals wereexposed onto M-1cells after48hours of coculture and quantification of adherent COMcrystals were evaluated. Expression analysis of kidney stone-and adipocytes-relatedinflammatory genes and proteins were performed.Results:Inflammatory including monocyte chemoattractant protein-1(MCP-1),osteopontin (OPN)and tumor necrosis factor-α (TNF-α) were up-regulated markedly in coculture groups.OPNover-expression of M-1occurs in coculture systems contained RAW264. TNF-α andMCP-1over-expression are detected from coculture systems contained3T3-L1. Suchinflammatory changes are induced by the coculture without direct contact, suggesting therole of soluble factors. Coculturing M-1s with3T3-L1s and RAW264s resulted insignificantly increased adhesion of calcium oxalate monohydrate (COM) crystals to M-1.Conclusion: Our results postulate that paracrine loop involving renal tubular cells,adipocytes and/or macrophages aggravates inflammatory changes of renal tubular cells inmetabolic syndrome or obesity, which may promote the susceptibility of urolithiasis. Part2：Inflammatory Changes and Effect ofAdiponectin on KidneyCrystal Formation in Metabolic Syndrome Model MiceObjective:Although an epidemiological link between the metabolic syndrome and kidney stoneformation has been reported, the mechanism by which metabolic syndrome promoteskidney stone formation has yet to be elucidated. We investigated calcium oxalate (CaOx)kidney stone formation in a mouse metabolic syndrome model.The aims of the presentstudy were to elucidate a possible mechanism of kidney crystal formation by using ametabolic syndrome (MetS) mouse model and to assess the effectiveness of adiponectintreatment for the prevention of kidney crystals.Methord:We induced CaOx crystal deposition in8-week-old male ob/ob mouse, and a control strain,Wild-type(+/+) mouse, by administering50mg/kg glyoxylate (GOx) for6days. Mice weredivided into3groups: Wild-type (+/+) mouse(control, n=12); ob/ob mouse (stone forming,n=12); APN treatment (n=12);Urine and blood samples were collected for biochemistrytesting, and the kidneys were harvested for estimation of crystal deposition anddeterminations of the expression of osteopontin(OPN),monocyte chemoattractantprotein-1(MCP-1) andAdiponectin(APN).Results:Wild-type (+/+) mice showed no kidney crystal formation, whereas ob/ob mice showedcrystal depositions in their renal tubules. However, this deposition was remarkably reducedby adiponectin.Significant upregulation of both inflammatory OPN and MCP-1was seenin the kidneys of ob/ob mice and whereas downregulation of antiinflammatory cytokineAPN. By TUNEL staining, ob/ob and ob/ob+APN kidneys showed stained nuclei in thearea from the cortex to medulla, with the cortico-medullary junction showing particularlystrong staining. Conclusion:The results of this study provide compelling evidence that the mechanism of kidney crystalformation in the MetS environment involves the progression of an inflammation, mightincluding oxidative stress and apoptosis in renal tissues. This is the first report to prove thepreventive effect of adiponectin treatment for kidney crystal formation by renoprotectiveactivities and inhibition of inflammation and apoptosis.