It was pointed out by many researches that keeping the concentration of Ca2+ in cells could increase the survival rate of hypothermic preserved kidneys and the survival rate of transplants. In this study, changes of the concentration of calcium were detected within catoplasm, mitochondria, endoplasmic reticulum and nucleus in the isolated hypothermic storage cat kidney, Ca2+ be marked with calcium cytochemical probe (K2H2Sb2O7) and detected by X-ray microanalysis of microsections. After 24, 48 and 72 hours preservation, the p/b (peak/back) of calcium within cytoplasm and mitochondria increased significantly. There were no obvious changes within endoplasmic reticulum and nucleus. It demonstrates that the Ca2+ were released from calcium pool (except the endoplasmic reticulum, nucleus, mitochondria etc.) to cytoplasm during preservation; and mitochondria can uptake calcium from cytoplasm to some extent, while the calcium concentration of cytoplasm is higher than normal.
Animals ; Calcium ; metabolism ; Cats ; Cryopreservation ; Female ; In Vitro Techniques ; Kidney Cortex ; cytology ; metabolism ; Male ; Spectrometry, X-Ray Emission ; Time Factors

OBJECTIVETo observe the effect of Cordyceps sinensis (CS) powder on renal oxidative stress and mitochondria functions in 5/6 nephrectomized rats, and to primarily explore its possible mechanisms.

METHODSTotally 30 male Sprague-Dawley rats were divided into the sham-operation group, the model group, and the treatment group by random digit table, 10 in each group. A chronic kidney disease (CKD) rat model was prepared by one step 5/6 nephrectomy. Rats in the treatment group were intragastrically administered with CS powder solution at the daily dose of 2 g/kg, once per day. Equal volume of double distilled water was intragastrically administered to rats in the sham-operation group and the model group. All medication lasted for 12 weeks. The general condition of rats, their body weight, blood pressure, 24 h proteinuria, urinary N-acetyl-β-D-glucosaminidase (NAG), serum creatinine (SCr) , and blood urea nitrogen (BUN) were assessed before surgery, at week 2, 4, 6, 8, 10, and 10 after surgery. Pathological changes of renal tissues were observed under light microscope. Morphological changes of mitochondria in renal tubular epithelial cells were observed under transmission electron microscope. Activities of antioxidant enzymes including reduced glutathione (GSH), manganese superoxide dismutase (MnSOD), and malondialdehyde (MDA) in fresh renal tissue homogenate were detected. Mitochondria of renal tissues were extracted to detect levels of mitochondrial membrane potential and changes of reactive oxygen species (ROS). And expressions of cytochrome-C (Cyto-C) and prohibitin in both mitochondria and cytoplasm of the renal cortex were also measured by Western blot.

RESULTS(1) Compared with the sham-operation group, body weight was significantly decreased at week 2 (P <0. 01), but blood pressure increased at week 4 (P <0. 05) in the model group. Compared with the model group, body weight was significantly increased at week 12 (P <0. 01), but blood pressure decreased at week 8 (P < 0. 01) in the treatment group. (2) Compared with the sham-operation group, 24 h proteinuria, urinary NAG, blood SCr and BUN significantly increased in the model group (all P <0. 01). Compared with the model group, blood and urinary biochemical indices all significantly decreased in the treatment group (all P <0. 01). (3) Results of pathological renal scoring: Glomerular sclerosis index, scoring for tubulointerstitial fibrosis, degree of tubulointerstitial inflammatory infiltration were all obviously higher in the model group than in the sham-operation group (all P <0. 01). All the aforesaid indices were more obviously improved in the treatment group than in the model group (all P <0. 01). (4) Compared with the sham-operation group, activities of MnSOD and GSH-Px were significantly reduced, but MDA contents obviously increased in the renal cortex of the model group (all P <0. 01). Compared with the model group, activities of MnSOD and GSH-Px obviously increased (P <0. 05, P <0. 01), but MDA contents obviously decreased in the renal cortex of the treatment group (P <0. 01). (5) Compared with the sham-operation group, the mitochondrial membrane potential significantly decreased, but ROS levels significantly increased in the model group (all P <0.01). Compared with the model group, mitochondrial transmembrane potential increased in the treatment group, thereby inhibiting the tendency of increased production of ROS (both P < 0. 01). (6) Results of Western blot showed that, compared with the sham-operation group, expression levels of mitochondrial Cyto-C and Prohibitin were significantly reduced in the renal cortex (P <0. 01), but significantly elevated in the cytoplasm of the model group (P <0. 01). Compared with the model group, each index was obviously improved in the treatment group with statistical difference (P <0. 05, P <0. 01).

CONCLUSIONCS powder had renal protection, and its mechanism might partially depend on in- hibition of oxidative stress and protection for mitochondria.

Acetylglucosaminidase ; metabolism ; Animals ; Blood Urea Nitrogen ; Cordyceps ; Drugs, Chinese Herbal ; pharmacology ; Kidney ; Kidney Cortex ; Kidney Diseases ; Kidney Function Tests ; Male ; Malondialdehyde ; metabolism ; Mitochondria ; Nephrectomy ; Oxidative Stress ; drug effects ; Proteinuria ; Rats ; Rats, Sprague-Dawley ; Superoxide Dismutase ; metabolism

OBJECTIVETo observe the effect of Chinese herbs for stasis removing and collaterals dredging (CHSRCD) upon angiotensin-converting enzyme 2-angiotensin-(1-7)-Mas axis in the renal cortex of diabetic nephropathy rats.

METHODSTotally 89 male Sprague-Dawley rats were randomly divided into the blank control group (C group, n=22), the high-glucose high-fat control group (H group, n=10), and the streptozotocin (STZ)-injecting group (n=57). The diabetes rat model (n=50) was induced by feeding high-glucose high-fat diet in combination with intraperitoneal injection of STZ, which were further divided into the model group (M group, n=24), the irbesartan group (I group, n=13), and the CHSRCD (Z group, n=13). Rats in I and Z groups were intragastrically fed with suspension of irbesartan and CHSRCD, once daily for 16 weeks. Equal volume of drinking water was administrated to rats in the rest groups. Blood glucose and 24 h urine protein quantitation were tested at four time points. And the mRNA expression of ACE2 and Mas at various time points was detected by Real-time PCR, immunohistochemical assay, and Western blot. Quantitative analyses of ACE2 and Mas protein expression were performed at the end of week 16.

RESULTSCompared with the C group, blood glucose increased in the H and M groups (P < 0.01). It was higher in the H group (P < 0. 01). 24 h urine protein quantitation at different time points increased in the M group, and it was higher than that in the H group (P < 0.05). Compared with the M group, 24 h urine protein quantitation decreased at the end of week 8 in the I group, and at the end of week 8 and 16 in the Z group (P < 0.05). It was lower in the Z group than in the I group at the end of week 16 (P < 0.05). Compared with the C and H groups, the expression of ACE2 mRNA in the renal cortex was lower in the M group at the end of week 16 (P < 0.01). Compared with the M group, it was higher in the Z group (P < 0. 01). There was no statistical difference in the expressions of Mas mRNA at the end of week 16 between the C group and the M group (P > 0.05). It was lower in the M group than in the H group (P < 0.05). It was higher in the Z group than in the M group (P < 0.05), and higher than in the I group (P < 0.05). The expression of ACE2 and Mas protein in the M group decreased as time went by. The expression quantitation of ACE2 and Mas protein at the end of week 16 was lower in the M group than in the C group (P < 0.05). Compared with the M group, ACE2 expression of the Z group and Mas of the I and Z groups increased more significantly (P < 0. 05).

CONCLUSIONCHSRCD could play a role in renal protection for diabetic nephropathy rats by up-regulating the mRNA and protein expression of ACE2 and Mas, promoting the ACE2-Ang-(1-7)-Mas axis, and lowering urinary protein.

Angiotensin I ; metabolism ; Animals ; Diabetes Mellitus, Experimental ; metabolism ; Diabetic Nephropathies ; metabolism ; Drugs, Chinese Herbal ; pharmacology ; Kidney Cortex ; metabolism ; Male ; Peptide Fragments ; metabolism ; Peptidyl-Dipeptidase A ; metabolism ; Proto-Oncogene Proteins ; metabolism ; Rats ; Rats, Sprague-Dawley ; Receptors, G-Protein-Coupled ; metabolism

OBJECTIVETo study the expression of TGF-beta(1)vimentin and desmin in the renal cortex of diabetic rats induced by STZ.

METHODSDiabetes was induced in 24 male SD rats by single intraperitoneal injection of 1.0%STZ (70 mg/kg). Twenty-four age, weight and sex matched SD rats were used as controls. The expression of TGF-beta(1),vimentin and desmin mRNA in the renal cortex were detected by RT-PCR on the 3rd, the 7th, the 14th and the 30th day after the DM rat model established.

RESULT(1)The expression of TGF-beta(1), vimentin mRNA in the renal cortex of diabetic rats gradually increased respectively from the 7th day and the 14th day after the model established, and the expressive intensity was significantly greater than that in controls (P<0.05 or P<0.01). However,the expression of desmin mRNA in the renal cortex of diabetic rats gradually decreased from the 14th day after the model established, and the expressive intensity was significantly less than that in controls (P<0. 05 or P<0.01). (2) The expression of TGF-beta(1)mRNA correlated positively to that of vimentin mRNA (r 0.740 P=0.000), while the expression of desmin mRNA correlated negatively to that of TGF-beta(1)mRNA (r 0.695 P=0.000) and to that of vimentin mRNA (r 0.591 P=0.002).

CONCLUSIONThe expression of renal cortical TGF-beta(1) and vimentin mRNA gradually increase while the expression of desmin mRNA gradually decrease during the first month of the diabetic model established suggest TGF-beta(1) may play a role in the transformation of renal tubular epithelial cells into fibroblast during the progressive interstitial fibrosis of diabetic nephropathy.

Animals ; Desmin ; genetics ; Diabetes Mellitus, Experimental ; metabolism ; Kidney Cortex ; metabolism ; Male ; Rats ; Rats, Sprague-Dawley ; Streptozocin ; Transforming Growth Factor beta ; genetics ; Transforming Growth Factor beta1 ; Vimentin ; genetics

Estimation of the postmortem interval (PMI) is a practical task in daily forensic casework. Researches on PMI is an important practical project in forensic field. Estimation of the time since death is influenced by internal and external, antemortem and postmortem factors, thus the old methods have limitations. Fourier transform infrared (FTIR) spectroscopy has been applied to study the pure protein, nucleic acid and carbohydrate and to detect the changes in complex cells and tissues. At present because the powerful software has could be used to achieve the spectrum transformation, smoothing, baseline correction and normalization, it is possible to analyze the samples quantitatively with the FTIR which has been applied in the biology and clinical medicine. This paper has reviewed the mechanism of FTIR and its application in biomedicine. The postmortem FTIR spectral changes were also discussed, which showed its potential for estimating PMI.
Animals ; Forensic Pathology/methods* ; Glycogen/biosynthesis* ; Humans ; Kidney Cortex/metabolism* ; Lung/metabolism* ; Muscle, Skeletal/metabolism* ; Myocardium/metabolism* ; Nucleic Acids/metabolism* ; Postmortem Changes ; Rats ; Spectroscopy, Fourier Transform Infrared/methods* ; Spleen/metabolism* ; Temperature ; Time Factors

Posttransplant diabetes mellitus, a complication due to corticosteroids and the calcineurin inhibitors, cyclosporine and tacrolimus, is commonly regarded as a form of type 2 diabetes mellitus. Diabetes ketoacidosis, which requires relative insulin deficiency to impair fatty acid metabolism, is a complication of type 1 diabetes mellitus. We report two patients who presented with diabetic ketoacidosis after kidney transplantation. Two patients presented with severe hyperglycemia, significant ketosis and metabolic acidosis of variable severity. One patient was treated with a cyclosporine-based regimen, and the other with a tacrolimus-based regimen. Both were found to have moderate to high serum levels of calcineurin inhibitors on presentation. Our experience suggests that post-transplant diabetes mellitus, in association with calcineurin inhibitor, may result in ketoacidosis either secondary to relative beta cell dysfunction, peripheral insulin resistance, or a combination of the two effects. Post transplant diabetes mellitus can be an atypical form of adult-onset diabetes with features of both type 1 and type 2 diabetes mellitus.
Acidosis ; Adrenal Cortex Hormones ; Calcineurin ; Cyclosporine ; Diabetes Mellitus ; Diabetes Mellitus, Type 1 ; Diabetes Mellitus, Type 2 ; Diabetic Ketoacidosis ; Humans ; Hyperglycemia ; Insulin ; Insulin Resistance ; Ketosis ; Kidney Transplantation ; Metabolism ; Tacrolimus

Posttransplant diabetes mellitus, a complication due to corticosteroids and the calcineurin inhibitors, cyclosporine and tacrolimus, is commonly regarded as a form of type 2 diabetes mellitus. Diabetes ketoacidosis, which requires relative insulin deficiency to impair fatty acid metabolism, is a complication of type 1 diabetes mellitus. We report two patients who presented with diabetic ketoacidosis after kidney transplantation. Two patients presented with severe hyperglycemia, significant ketosis and metabolic acidosis of variable severity. One patient was treated with a cyclosporine-based regimen, and the other with a tacrolimus-based regimen. Both were found to have moderate to high serum levels of calcineurin inhibitors on presentation. Our experience suggests that post-transplant diabetes mellitus, in association with calcineurin inhibitor, may result in ketoacidosis either secondary to relative beta cell dysfunction, peripheral insulin resistance, or a combination of the two effects. Post transplant diabetes mellitus can be an atypical form of adult-onset diabetes with features of both type 1 and type 2 diabetes mellitus.
Acidosis ; Adrenal Cortex Hormones ; Calcineurin ; Cyclosporine ; Diabetes Mellitus ; Diabetes Mellitus, Type 1 ; Diabetes Mellitus, Type 2 ; Diabetic Ketoacidosis ; Humans ; Hyperglycemia ; Insulin ; Insulin Resistance ; Ketosis ; Kidney Transplantation ; Metabolism ; Tacrolimus

OBJECTIVESTo explore the effects and significance of Huanshuai Recipe Oral Liquid (, HSR), a formula with supplementing qi, nourishing blood and activating blood on restructuring glomerular microvasculature and expression of vascular endothelial growth factor (VEGF) in subtotal nephrectomized (SNX) rats.

METHODSA total of 76 male Wistar rats were randomly divided into four groups: 16 in the sham-operated group and fed with tap water 10 mL/kg per day; 20 in the model group were operated with 5/6 SNX and fed with tap water 10 mL/ kg per day; 20 SNX rats in the HSR group were treated with HSR 10 mL/kg per day; 20 SNX rats in the losartan group were treated with losartan 40 mg/kg per day. Serum creatinine (SCr) and urinary protein excretion (Upro) were examined at the 2nd, 4th, 8th, and 12th weeks of the treatment, and the remnant kidneys were harvested. Changes in histological microstructure were evaluated using light microscopy, and the expression of VEGF was detected by using ELISA.

RESULTSUpro, microvasculature injury and glomerulosclerosis were found to be alleviated in HSR and Losartan groups, respectively. The change of VEGF expression showed positive correlation with glomerular capillary area and peritubular capillary number (r=0.448, r=0.422, P<0.01), but negative correlation with that of SCr and Upro (r=-0.592, r=-0.481, P<0.01).

CONCLUSIONSHSR could regulate the VEGF expression, reduce the loss of microvasculature, which demonstrated similar renal protective effects to losartan in SNX rats. Examination of Chinese herbal medicine influence on VEGF signaling and restructuring renal microvasculature may elucidate the molecular mechanism of renal protection to a certain degree.

Administration, Oral ; Animals ; Capillaries ; drug effects ; metabolism ; pathology ; Collagen Type IV ; metabolism ; Creatinine ; blood ; Drugs, Chinese Herbal ; administration & dosage ; pharmacology ; Extracellular Matrix ; drug effects ; metabolism ; Fibronectins ; metabolism ; Immunohistochemistry ; Kidney Cortex ; drug effects ; metabolism ; pathology ; Kidney Function Tests ; Kidney Glomerulus ; blood supply ; drug effects ; pathology ; physiopathology ; Microvessels ; drug effects ; Nephrectomy ; Proteinuria ; blood ; drug therapy ; physiopathology ; Rats ; Rats, Wistar ; Time Factors ; Vascular Endothelial Growth Factor A ; metabolism

OBJECTIVETo investigate the expressions of 78-kDa glucose-regulated protein (GRP78) and Caspase-12 and their relationship with apoptosis in renal cortex of diabetic rats.

METHODSUninephrectomized Wistar rats were used to induce diabetes by intraperitoneal injection of Streptozotocin (STZ 65 mg/kg). After 8 weeks, the expression and distribution of GRP78, Caspase-12, proliferating cell nuclear antigen (PCNA) were examined by immunohistochemistry. Flow cytometry was used to detect the levels of protein of GRP78 and Caspase-12. Apoptosis was evaluated by means of terminal deoxynucleotidyl transferase-mediated d-UDP nick-end labeling (TUNEL) and Flow cytometry. Serum creatinine, blood urea nitrogen and 24-hour urine protein excretion were checked.

RESULTSCompared with those in normal control group, the numbers of apoptosis and the expression of GRP78, Caspase-12 in glomerular and tubular cells were much higher in the diabetic kidneys at 8 weeks. There was no significant difference between group A and group B.

CONCLUSIONActivation of endoplasmic reticulum stress may play an important role in the development of diabetic nephropathy.

Animals ; Apoptosis ; Caspase 12 ; metabolism ; Diabetes Mellitus, Experimental ; complications ; Diabetic Nephropathies ; pathology ; Endoplasmic Reticulum Stress ; Heat-Shock Proteins ; metabolism ; Kidney Cortex ; metabolism ; pathology ; Male ; Proliferating Cell Nuclear Antigen ; metabolism ; Rats ; Rats, Wistar

OBJECTIVETo investigate the effect of hyperglycemia and pioglitazone on TGF-beta(1) gene expression in peripheral blood mononuclear cells (PBMC) and renal cortex, and the correlation of TGF-beta(1)mRNA levels between PBMC and renal cortex in STZ induced diabetic rats.

METHODSFifty-four Sprague-Dawley rats were randomly divided into three groups: 18 normal control rats (group C), 18 diabetic rats (group D) and 18 diabetic rats treated with pioglitazone (20 mg x kg(-1)x d(-1), group DP). Six rats from each group were sacrificed at 2, 4, 8 weeks. TGF-beta(1)mRNA levels of PBMC and renal cortex were examined by RT-PCR+Slit hybridization analysis.

RESULTTGF-beta(1)mRNA level of renal cortex in group D was significantly higher than that in group C at each time point (P<0. 05); TGF-beta(1)mRNA level of PMBC in group D was slightly higher than that of group C at 4 weeks, and significantly higher at 8 weeks (P=0.01). There was positive correlation of TGF -beta(1)mRNA level between PBMC and renal cortex before (r=0.83, P=0.02) and after pioglitazone treatment at 8 weeks (r=0.82, P=0.03).

CONCLUSIONTGF-beta(1)mRNA level of PBMC may reflect the change of TGF-beta(1) gene expression of renal cortex in diabetic rats.

Animals ; Diabetes Mellitus, Experimental ; metabolism ; Female ; Kidney Cortex ; metabolism ; Leukocytes, Mononuclear ; metabolism ; Male ; RNA, Messenger ; analysis ; Rats ; Rats, Sprague-Dawley ; Streptozocin ; Transforming Growth Factor beta ; genetics ; Transforming Growth Factor beta1