International Cooperation ; Nephrology ; trends ; Pediatrics ; trends

Objective To explore the molecular effect and interaction among nephrin, podocin, CD2AP and ?-actinin-4. Methods Firstly, the recombinant RNA interference (RNAi) plasmid-psiRNA-hH1GFPzeo, specifically targeting to the mRNA of nephrin, podocin, CD2AP or ?-actinin-4, was respectively tansfected into the mouse podocyte clone (MPC5) to each knockdown (KD) the expression of nephrin, podocin, CD2AP or ?-actinin-4. Molecular distributions were revealed by confocal microscopy, and the mRNA and protein expressions were detected with semi-quantitative RT-PCR and Western blotting. Results (1)In podocin KD group (siPod966 and siPod54), the mRNAs of podocin and nephrin were not detected, their protein decreased 92% and 79%, 82% and 67%, respectively. The mRNA and protein level of CD2AP increased 62% and 42%, 71% and 46%, respectively, whereas ?-actinin-4 did not change. In nephrin KD group (siNep492), the mRNA expression and protein level of nephrin were not detected, CD2AP increased 35% and 48%, respectively; and whereas podocin and ?-actinin-4 did not change. In CD2AP KD group (siCda744 and siCda21), the mRNA of expression CD2AP was not detected, and its protein level decreased 92% and 83%, the mRNA and protein of nephrin decreased 60% and 48%, 76% and 72%, respectively; podocin increased 38% and 22%, 56% and 44%, respectively; whereas ?-actinin-4 did not change. In ?-actinin-4 KD group (siAct1790 and siAct319), the mRNAs expression of ?-actinin-4 and nephrin decreased 69% and 58%, 64% and 49%, respectively; their protein level decreased 81% and 55%, 71% and 64%, respectively. However, the mRNAs of podocin and CD2AP increased 50% and 34%, 45% and 28%, respectively; and their protein level increased 64% and 46%, 65% and 42%, respectively. (2) With their expression change, the distributions of nephrin, podocin and CD2AP shifted evidently from the cell membrane surface to the nucleus circumference, whereas ?-actinin-4 showed no change, which was still localized in the cytoplasm and further extended to foot processes. Conclusion (1) Nephrin might more independently play a crucial role in the slit diaphragm complex. (2) Alpha-actinin-4 might interact direcdy or indirectly with nephrin, podocin and CD2AP. (3) The relationship among these podocyte molecules might not be spontaneous, either a single-directional or bi-directional reaction. (4) The normal localization of these podocyte molecules might depend on their normal expression quantity.

Objective To dynamically observe the expression of slit diaphragm complex molecules, including nephrin, podocin, CD2AP, and cytoskeleton protein a-actinin-4, in adriamycin-induced nephrotic (ADN) rats, and to further explore the molecular behavior of podocyte proteins during the occurrence and development of proteinuria and their possible mechanisms. Methods Adriamycin nephropathy was induced by a single tail intravenous injection of adriamycin. Renal tissue samples were collected at day 3, 7, 14, and 28, respectively. The distribution, mRNA expression and protein expression of nephrin, podocin, CD2AP and a-actinin-4 were examined by indirect immunofluorescence, real-time PCR and Western blotting, respectively. Results (1) After the adriamycin injection, a significant increment of the 24-hour urinary protein was observed at day 14 and persisted up to day 28 (P

Objective To explore the molecular mechanisms underlying therapeutic responses of the anti-proteinuria drugs from the view of podocyte molecule. Methods Adriamycin (ADR) nephropathy was induced by a single tail intravenous injection of adriamycin. Lisinopril, prednisone and all-trans retinoic acid (ATRA) were administered once a day to the adriamycin-induced nephrotic rats at the first day after adriamycin injection respectively. Renal tissue samples were collected at day 3, 7, 14, and 28 after adriamycin injection respectively. The distribution, mRNA expression and protein expression of nephrin, podocin, CD2AP and ?-actinin-4 were examined by indirect immunofluorescence, real-time PCR and Western blotting, respectively. The interactions among nephrin and podocin, nephrin and CD2AP, as well as the nephrin phosphorylation were detected by immunoprecipitation, respectively. Results Compared to the control rats, 24 h urinary protein of the ADR rats increased significantly at day 14 (P

BACKGROUND:Studies on high glucose exposure in human periodontal ligament cel s usual y focus on the biological behaviors, pathways and secretory factors, but whether the metabolic memory is involved is little known. OBJECTIVE:To investigate the metabolic memory of high glucose exposure in human periodontal ligament cel s. METHODS:Human periodontal ligament cel s were primarily cultured and identified. Cel s at 5-8 passages were selected and randomized into four groups. Group A (controls):DMEM containing 5.5 mmol/L glucose for 8 days;group B (5-day memory group):DMEM containing 35 mmol/L glucose for 3 days and DMEM containing 5.5 mmol/L glucose for 5 days;group C (3-day memory group):DMEM containing 35 mmol/L glucose for 5 days and DMEM containing 5.5 mmol/L glucose for 3 days;group D (8-day high glucose group):DMEM containing 35 mmol/L glucose for 8 days. The cel proliferation was detected by cel counting kit-8, the cel apoptosis was determined by flow cytometry, and the levels of total proteins and alkaline phosphatase were investigated using ELISA. RESULTS AND CONCLUSION:Compared with the control group, the cel proliferation in the other three groups was significantly reduced (P<0.05), the number of apoptotic cel s was significantly increased, while the levels of total proteins and alkaline phosphatase were significantly decreased (P<0.05). These results suggest that high glucose causes persistent changes in human periodontal ligament cel s by inhibiting cel viability, increasing the apoptosis and downregulating the levels of the total proteins and alkaline phosphatase

0.05) at any observed time points as compared with the controls.3.From day 7 after the adriamycin injection,VEGF protein reduced significantly(P

OBJECTIVETo explore the inhibition of Tangtong Recipe (TTR) on proliferation of vascular smooth muscle cells (VSMCs) in rat model of diabetes mellitus using seropharmacological and modern molecular biological technologies.

METHODSThe drug-serum of TTR was prepared by feeding pure-breed New Zealand rabbits at various doses (18 mL/kg, 13 mL/kg, and 8 mL/kg) of TTR, and used to intervene the VSMCs get from the iliac artery of diabetic model rats cultured by tissue block plantation. The impact of drug-serum on VSMCs proliferation was estimated through measuring tritiated thymine deoxy-ribonucleotide (3H-TDR) incorporation using liquid-scintillation detector, and the cell cycle was detected using flow cytometry.

RESULTSThe 3H-TDR incorporation in cells after intervention with various doses of TTR drug-serum were reduced significantly, in a concentration-dependent manner, to the level lower than that in the normal serum control group (P < 0.01). And the percentage of S and G2/M phase cells in the drug-serum treated groups also were markedly lower than that in the normal control group (P < 0.01).

CONCLUSIONDrug-serum of TTR could inhibit the proliferation of VSMC of diabetic rat in vitro in a concentration-dependent manner.

Animals ; Cell Proliferation ; drug effects ; Cells, Cultured ; Diabetes Mellitus, Experimental ; pathology ; Dose-Response Relationship, Drug ; Drugs, Chinese Herbal ; pharmacology ; Iliac Artery ; cytology ; Male ; Muscle, Smooth, Vascular ; pathology ; Rabbits ; Random Allocation ; Rats ; Rats, Wistar ; Serum

OBJECTIVETo evaluate the intervention effect of diet, exercise and Jiangtang Bushen Recipe (JBR, a Chinese herbal recipe) in preventing the progress of patients with impaired glucose tolerance (IGT) to diabetes mellitus (DM) type 2.

METHODSFifty-one patients with IGT, with their diagnosis conformed to the diagnosis standard of WHO, 1999, were randomly divided into the control group (n = 26) and the TCM group (n = 25). Patients in the control group attended to the educational course for DM and received dietotherapy and kinetotherapy, and to those in the TCM group, under these treatments, JBR was given additionally. Oral glucose tolerance test (OGTT), body weight index (BWI), levels of blood lipids and fasting insulin of all the patients were examined after 3 months, 6 months and 12 months of treatment. The total observation time was 1 year.

RESULTSExcept the 6 cases out of the 51 patients (11.7%), on whom the observation discontinued, in the control group, as compared with before treatment, levels of fasting insulin and fasting blood glucose after treatment were not changed significantly (P > 0.05), also insignificant difference was shown in levels of total cholesterol (TC) and triglyceride (TG), though the two indexes lowered slightly after treatment (P > 0.05), but significant difference was shown in comparison of OGTT/2 h, blood glucose and BWI (P < 0.05). While in the TCM group, fasting blood glucose was changed insignificantly (P > 0.05), but there was significant difference in comparison of fasting insulin, TC, BWI, OGTT/2 h and plasma glucose levels (P < 0.01) respectively before and after treatment. At the end of the study, the cumulative cases with conversion to diabetes were 3 (13.6%) in the control group, and 1 (4.3%) in the TCM group, chi 2 test showed insignificant difference in comparison of diabetes conversion rate between the two groups (P > 0.05), however, the TCM group showed a better year conversion rate of normal glucose tolerance than that in the control group (chi 2 = 8.31, P < 0.01).

CONCLUSIONTCM intervention is possibly effective in delaying the conversion of IGT to DM type 2, and plays integrative effeciency in impelling IGT patients to health. The favorable education and treatment of DM controlling, including dieto- and kineto-therapy may also be advantageous in IGT intervention, but could not be effective in blocking the advance of IGT.

Adult ; Aged ; Combined Modality Therapy ; Diabetes Mellitus, Type 2 ; prevention & control ; Drugs, Chinese Herbal ; therapeutic use ; Exercise ; Female ; Follow-Up Studies ; Glucose Intolerance ; drug therapy ; Glucose Tolerance Test ; Humans ; Male ; Middle Aged ; Phytotherapy

The combination of Glycyrrhizae Radix et Rhizoma and Aconiti Lateralis Radix Preparata can increase efficacy and decrease toxicity. This study started from the phenomena of protein self-assembly in the mixed decoction of Glycyrrhizae Radix et Rhizoma with Aconiti Lateralis Radix Preparata. The attenuated mechanism was explored between the combination of Glycyrrhizae Radix et Rhizoma and Aconiti Lateralis Radix Preparata by using the protein of Glycyrrhizae Radix et Rhizoma and aconitine which was the major toxic component of Aconiti Lateralis Radix Preparata. Glycyrrhizae Radix et Rhizoma protein with aconitine could form stable particles which particle mean diameter was (206.2 ± 2.02) nm and (238.20 ± 1.23) nm at pH 5.0 in normal temperature. Through the mouse acute toxicity experiment found that injection of aconitine monomer all mice were killed, and injection of Glycyrrhizae Radix et Rhizoma protein-aconitine particles with the same content of aconitine all mice survived. Survey the stability of Glycyrrhizae Radix et Rhizoma protein-aconitine shows that the colloid particles is stable at room temperature, and it has the possibility to candidate drug carrier. Glycyrrhizae Radix et Rhizoma protein can reduce the toxicity of aconitine through self-assembly.
Aconitum ; chemistry ; toxicity ; Animals ; Drugs, Chinese Herbal ; toxicity ; Female ; Glycyrrhiza ; chemistry ; toxicity ; Male ; Mice ; Mice, Inbred ICR ; Plant Proteins ; chemistry ; isolation & purification ; toxicity ; Rhizome ; chemistry ; toxicity

OBJECTIVE: To understand WT1 mutations in patients with steroid resistant nephrotic syndrome (SRNS) accompanied with genitourinary malformations. METHODS: Three cases of SRNS accompanied with genitourinary malformations were enrolled. The expression of podocyte molecules (nephrin, podocin, alpha-actinin-4, WT1, and CD2AP) in 2 cases was analyzed with the immunofluorescence and immunohistochemistry techniques. The genomic DNA and cDNA of WT1 were analyzed by using PCR and RT-PCR, respectively. GeneScan and GeneScan software were used to quantify the ratio of +KTS/-KTS isoforms. RESULTS: The onset ages of 3 cases were 6 months, 1 year, and 10 years old, respectively. The diagnosis age was 7 months, 9 years, and 15 years old, respectively. The phenotype of Case 1 and Case 3 was male accompanied with genitourinary malformations. Case 2 was phenotypic female. Karyotype analysis of 3 cases revealed 46, XY. Three cases were diagnosed as SRNS. Focal segmental glomerulosclerosis (FSGS) was confirmed in 2 cases. Podocyte molecular expression altered in renal tissues of 2 cases. In addition, WT1 staining was negative in Case 1. WT1 expression in Case 2 showed diffuse nuclear staining with less obvious speckles compared with controls. WT1 IVS 9 +5 G>A mutation was detected in Case 2 and WT1 Exon 9 1186 G>A mutation was detected in Case 3. No WT1 mutation was detected in Case 1. CONCLUSION Karyotype analysis and WT1 genetic testing should be done in all female patients with early onset steroid resistant FSGS and in male patients with SRNS accompanied with genitourinary malformations. Abnormal podocyte molecular expression suggests that more podocyte molecules might be involved in the pathogenesis of proteinuria in WT1 mutational patients.
Adolescent ; Child ; Drug Resistance ; genetics ; Exons ; Female ; Hormones ; pharmacology ; Humans ; Infant ; Male ; Mutation ; Nephrotic Syndrome ; complications ; genetics ; Phenotype ; Urogenital Abnormalities ; complications ; genetics ; WT1 Proteins ; genetics