OBJECTIVETo explore the regulatory effect of CpG methyltransferase (M.SssI) on expression of claudin-7 and claudin-8, promoting apoptosis and inhibiting proliferation of human colorectal cancer HT-29 cells.

METHODSHT-29 cells were treated with M.SssI (50 U/ml) for 24 hours. The methylation status of claudin-7 and claudin-8 gene promoters was assayed by bisulfite sequencing PCR (BSP). Real-time PCR with SYBR green I technique was used to detect the relative expression of claudin-7 and -8 mRNA, and claudin-7 and claudin-8 proteins were tested by cell immunofluorescence and Western blotting, while the effect on cell apoptosis was assessed by Hoechst 33342 fluorescence and flow cytometry. Inhibition of cell proliferation was measured by MTT assay.

RESULTSThe amounts of methylated claudin-7 and claudin-8 gene CpGs were 25, 10 in the M.SssI group, 9 and 5 in the PBS group, 0 and 3 in the 5-azacytidine group, respectively. Compared with the PBS group, Claudin-7 and -8 were significantly reduced by M.SssI (P < 0.05), but increased by 5-azacytidine (P < 0.05) at both mRNA and protein levels. Hoechst 33342 staining revealed that HT-29 cells treated with PBS and 5-azacytidine were not significantly different, showing even blue fluorescence, round shape and same cell volume. But the M.SssI group presented more apoptotic cells with intensive white fluorescence intensity. Cytometry indicated that early apoptotic index of the M.SssI group was increased by 84.7%, compared with that of the PBS group (P = 0.002). Measurement of MTT optical density demonstrated that cell growth of the M.SssI group was significantly lower than that of the PBS group (P = 0.002), with an inhibition rate of 32.1%, whereas the proliferation of 5-azacytidine group was similar to that of the PBS group (P = 0.084).

CONCLUSIONSOur findings suggest that M.SssI can down-regulate claudin-7, -8 mRNA and proteins in the human colon cancer HT-29 cells by up-regulating methylation status of claudin-7 and -8 gene promoters, and finally induce apoptosis and inhibit proliferation of the tumor cells.

Apoptosis ; physiology ; Cell Proliferation ; Claudins ; metabolism ; Colonic Neoplasms ; DNA-Cytosine Methylases ; metabolism ; Down-Regulation ; physiology ; Flow Cytometry ; Gene Expression Regulation ; HT29 Cells ; Humans ; RNA, Messenger ; Real-Time Polymerase Chain Reaction

INTRODUCTIONA complex relationship exists between chronic kidney disease-mineral and bone disorder (CKD-MBD) and adverse outcomes among dialysis patients. This study aimed to report the prevalence of CKD-MBD and examine the impact of achieving target CKD-MBD parameters on morbidity and mortality one year after peritoneal dialysis (PD) initiation.

METHODSIn this retrospective cohort study, patients electively initiated on PD were followed up for one year. Laboratory parameters were collected and the prevalence of CKD-MBD 4-6 months after PD initiation was determined based on the Kidney Disease Outcomes Quality Initiative (KDOQI) and Kidney Disease: Improving Global Outcomes (KDIGO) guidelines. Linear regression and Cox proportional hazards model were used to evaluate the effects of achieving CKD-MBD targets 4-6 months after PD initiation on hospitalisation, the incidence of peritonitis or exit-site infections (ESIs), and mortality at one year.

RESULTSThe prevalence of CKD-MBD among the 86 patients in this study was 86.0% (KDOQI) and 54.7% (KDIGO). There was no significant difference in hospitalisation duration between patients who achieved targets and those who did not. Patients who failed to meet all the KDIGO CKD-MBD or calcium serum targets had a higher incidence of peritonitis or ESI. A trend toward shorter time to death was observed among patients who failed to meet the KDIGO phosphorus serum targets.

CONCLUSIONThere was moderate (KDIGO) to high prevalence (KDOQI) of CKD-MBD among the patients. Achievement of all the KDIGO CKD-MBD or calcium serum targets was associated with reduced peritonitis or ESI, while achievement of the KDIGO phosphorus serum targets was associated with improved survival.

Adult ; Aged ; Aged, 80 and over ; Chronic Kidney Disease-Mineral and Bone Disorder ; complications ; epidemiology ; Female ; Humans ; Kidney Failure, Chronic ; complications ; therapy ; Linear Models ; Male ; Middle Aged ; Peritoneal Dialysis ; Prevalence ; Proportional Hazards Models ; Quality Control ; Retrospective Studies ; Treatment Outcome

Animals ; Apoptosis ; Brain/metabolism* ; Male ; Phosphatidylinositol 3-Kinases/metabolism* ; Rats ; Rats, Sprague-Dawley ; Subarachnoid Hemorrhage/drug therapy* ; Thyroxine/pharmacology*