AIM: To investigate the mechanism of renal damage in chronic intermittent hypoxia (CIH) rat model.METHODS:The Sprague-Dawley rats were randomly divided into 2-week CIH group (2IH), 2-week simulated air control group (2C), 4-week CIH group (4IH) and 4-week simulated air control group (4C).HE staining, PAS staining and Masson staining were used for histological evaluation .Blood was collected for the measurement of superoxide dismutase (SOD).The mRNA expression of hypoxia-inducible factor-1α(HIF-1α), manganese superoxide dismutase (MnSOD), copper/zinc superoxide dismutase ( Cu/ZnSOD ) was detected by real-time PCR.RESULTS: ( 1 ) No significance difference of renal weight , body weight , and the ratio of renal weight to body weight was observed , while IH caused mor-phologic kidney damage , especially in 4IH group.Hypertrophy of epithelial cells in the kidney tubles and dilation in the glomeruli were observed under light microscope with HE and PAS staining , especially in 4IH group.Masson staining showed no significant fibrotic response in the kidney of the rats exposed to IH .(2) The SOD levels in the serum and kid-ney were decreased after CIH .Compared with the corresponding control groups , the levels of serum SOD were significantly lower in CIH groups, especially in 4IH group.The mRNA expression of Cu/ZnSOD and MnSOD in CIH groups decreased significantly as compared with control groups .The mRNA levels of HIF-1αwere significantly higher in CIH groups than those in the corresponding control groups .CONCLUSION: CIH induces abnormalities of glomeruli and convoluted tu-bules, while 4-week IH exposure has not led to fibrotic response .CIH participates in the process of renal oxidative stress damage by upregulating HIF-1αtranscription and downregulating Cu/ZnSOD and MnSOD transcription .

OBJECTIVEHuman herpesvirus 6 (HHV-6) isolates are classified into two variants, HHV-6A and HHV-6B, based on distinct genetic, antigenic and biological characteristics. HHV-6 has been associated with encephalitis in children recently. This study aimed to establish a real time PCR assay for simultaneous detection of the two subtypes of HHV-6, and apply this new assay to children with suspected encephalitis, then analyze the relationship between the infection with HHV-6 and encephalitis in children.

METHODThe universal primers and variant-specific TaqMan probes were designed based on the highly conserved sequences of the DNA polymerase gene (U38) of HHV-6. The 5' end of the probes for HHV-6A and HHV-6B was labeled with the fluorescein reporter tetrachloro-6-carboxyfluorescein and 6-carboxyfluorescein (6-FAM), separately, while the 3' end were quenched with 6-carboxy-tetramethylrhodamine. The real time PCR assay for simultaneous detection of HHV-6A and HHV-6B was established. Then, the plasmids of HHV-6A and -6B which were diluted by a 10-fold series from 10(9) to 10(0) copies/microl, together with controls were used for testing both sensitivity and specificity of the real time PCR assay. The cerebrospinal fluid (CSF) specimens from 445 cases of suspected encephalitis were tested with this real time PCR and positive samples were then sequenced.

RESULTBoth HHV-6A (strain ZJ-159) and HHV-6B (strain GS) were positive on the real time PCR assay. There were no cross-reaction with herpes simplex virus type 1, type 2 (HSV-1, HSV-2), varicella-zoster virus (VZV), cytomegalovirus (CMV), Epstein-Barr virus (EBV), hepatitis B virus, Staphylococcus aureus, Mycoplasma pneumoniae and human DNA. A linear regression curve was obtained when plotting Ct values against the log10 of the viral DNA input for both subtypes of HHV-6. The sensitivity threshold was 10 copies/microl for the real time PCR. HHV-6 positive rate by the real time PCR assay was 4.72% (21/445), including 4 cases with HHV-6A infection, 16 cases of HHV-6B infection and 1 case with mixed HHV-6A and HHV-6B infection. The new PCR assay usually took 2 to 3 hours to provide results.

CONCLUSIONThis new real time PCR assay can simultaneously detect both subtypes of HHV-6, and have high specificity and sensitivity. It will provide an early and sensitive diagnosis of HHV-6 encephalitis in children.

Adolescent ; Child ; Child, Preschool ; DNA Fingerprinting ; DNA Primers ; DNA, Viral ; Encephalitis, Viral ; cerebrospinal fluid ; diagnosis ; virology ; Female ; Fluorometry ; Genotype ; Herpesvirus 6, Human ; genetics ; isolation & purification ; Humans ; Infant ; Infant, Newborn ; Polymerase Chain Reaction ; methods ; Sensitivity and Specificity

Abstract?AIM: To investigate mechanism of bradykinin ( BK) on inflammations of retinal pigment epithelium ( RPE) cells.?METHODS: ARPE -19 cells were cultured in vitro, stimulated by 100nM BK for 24h. Cell morphology changes were observed by microscope, and BK receptor localization was detected through cell immunofluorescence. Changes of Ca2+in BK and BR antagonist stimuli were detected by laser scanning confocal microscopy.The expressions of COX-1, COX-2, eNOS and iNOS protein in control group and BK group were detected by Western Blot.?RESULTS: After the stimulation of BK, there was no significant changes of ARPE-19 cells in morphology.Kinin B1 receptors ( B1R ) and B2 receptors ( B2R ) could be detected in ARPE-19 cells.Compared with control group, Ca2+concentrations significantly increased in BK group; in B1R antagonist group and B2R antagonist group Ca2+concentrations increased less than BK group; B1R and B2R antagonist group showed no obvious changes in Ca2+concentrations.Compared with control group, COX-2 and iNOS protein concentrations were significantly increased in BK group (P<0.001).?CONCLUSION:BK induces the increasing expression of COX-2 and iNOS in the cultured ARPE cells through binding with either B1R or B2R.

Zebrafish was selected as model animal, and glucocorticoid dexamethasone was used as a model compound to establish a rapid and high efficient osteopenia model. Zebrafish larvae at 4 days post fertilization (dpf) were exposed to a serial concentrations of dexamethasone solutions, and 0.5% DMSO was selected as the vehicle control group. All groups were incubated in 24-well plates (28.5 degrees C) until 9 dpf. In addition, effects of 10 micromol x L(-1) dexamethasone on preventing against osteopenia induced by etidronate disodium were also investigated. Zebrafish bones at 9 dpf were stained with alizarin red. Quantitative analysis of the stained area was performed by microscopic inspection and digital imaging methods to reflect the amount of bone mineralization. Results showed that dexamethasone group at 2.5, 10 and 25 micromol x L(-1) can decrease the staining area and the staining optical density values of zebrafish head bones when compared with the vehicle control group (0.5% DMSO), which suggested that dexamethasone can significantly reduce the zebrafish mineralized bone and the bone mineral density. Results also showed that 15 and 30 microg x mL(-1) etidronate disodium can increase the mineralized matrix of zebrafish head bone and prevent against osteopenia induced by dexamethasone. In conclusion, the study indicated that zebrafish can be an idea osteopenia model induced by dexamethasone.
Animals ; Bone Density ; drug effects ; Bone Density Conservation Agents ; pharmacology ; therapeutic use ; Bone Diseases, Metabolic ; chemically induced ; prevention & control ; Calcification, Physiologic ; drug effects ; Dexamethasone ; toxicity ; Disease Models, Animal ; Etidronic Acid ; pharmacology ; therapeutic use ; Larva ; drug effects ; growth & development ; Zebrafish

OBJECTIVETo investigate the influence of intrauterine infection caused by lipopolysaccharide on Toll-like receptor 4 (TLR4) signaling pathway in fetal and neonatal rat lungs in order to explore immunomodulating activity of innate immunity responding to intrauterine infection and its effect on lung development.

METHODSOn day 17 of pregnancy, 30 pregnant Sprague-Dawley (SD) rats were randomly divided into two groups: LPS group and saline group. For LPS group, LPS (10 microl, 40 microg/ml) was intrauterine injected between every two embryonic sacs of the pregnant rats, while the rats in the control group were injected with the same volume of pyrogen-free saline. Lung tissues of fetal rats and corresponding placental tissues were collected on the embryonic day 18 (E18), E20, and E22. Neonatal lung tissues were also harvested on postnatal day 1 (P1), P3, and P7. Lung sections and placental tissues were stained with hematoxylin and eosin for histological examination. Reverse transcription quantitative polymerase chain reaction (RT-PCR) analysis was performed to test mRNA expression for TLR4, myeloid differentiation 88 (MyD88) and IL-1beta, while immunohistochemistry was used to evaluate TLR4 and MyD88 expression in lung tissues. All data were analyzed with one-way analysis of variance (ANOVA) and q test.

RESULTS(1) Placental hematoxylin-eosin staining showed a great number of neutrophils infiltration, obvious interstitial hyperplasia and narrow capillaries in placental tissues in the LPS group which indicated that intrauterine infection occurred. However, there were no obvious inflammatory cells in the control group. (2) On E18, E20 and E22, the lung of LPS group showed no obvious pathological changes, and there were no apparent neutrophils infiltrated in alveoli, then some structural changes appeared. On P7, we found that the number of alveoli decreased, space of alveoli was larger than ever, septa thickened, but without significant constructive disorder. (3) In the LPS group, the TLR4, MyD88 and IL-1beta mRNA levels increased compared with control group, higher than control group at E20 and E22 (P < 0.05), and peaked at E22. Then the expression levels of these substances decreased slowly. (4) The result of immunohistochemistry showed that in lung tissues of the two groups at E18, there was no remarkable positive staining of TLR4 and MyD88, which mainly expressed in cytoplasm of bronchiole and alveolar epithelial cells, then positive cells increased slowly.

CONCLUSION(1) For perinatal rat lungs, intrauterine LPS infusion can induce an increased expression levels of TLR4 and MyD88 to a certain extent, which then returned to normal level gradually. At the same time, lung tissues showed a mild pathological change and inflammatory reaction. We propose that innate immune system of fetal lungs controls the magnitude of the LPS-induced cytokine response during the perinatal period. (2) The findings confirmed that LPS-activated signaling transduction pathway was the MyD88-dependent pathway.

Animals ; Animals, Newborn ; Female ; Infection ; metabolism ; Interleukin-1beta ; metabolism ; Lipopolysaccharides ; administration & dosage ; pharmacology ; Lung ; growth & development ; metabolism ; Myeloid Differentiation Factor 88 ; metabolism ; Pregnancy ; Pregnancy Complications, Infectious ; metabolism ; Prenatal Exposure Delayed Effects ; Rats ; Rats, Sprague-Dawley ; Signal Transduction ; drug effects ; Toll-Like Receptor 4 ; metabolism

Objective To analyze the HbA1c results of double heterozygotes of hemoglobin(Hb) NewYork and β-thalassemia detected by five different HbA1c detection systems,and compare the early-warning abilities of erroneous glycosylated hemoglobin results for Hb NewYork and β-thalassemia heterozygotes.Methods Peripheral blood samples from 40 patients without hemoglobinopathies with different range of HbA1c levels were collected to evaluate the consistency of the detected results.Variant Ⅱ system was used as the reference system which successfully completed NGSP Level Ⅰ laboratory certification.Variant Ⅱ Turbo 2.0 (V Ⅱ-T),Capillary 2 Flex Piercing(2FP),Primus Ultra2 (Ultra2)and Roche Modular PPI(PPI) 800 were used as the comparative systems.The HbA1 c in 2 sera from the patients with compand heterozygotes of Hb NewYork was detected by the above systems.Hemoglobin electrophoresis was performed.Gentypes of á-and β-globin genes were analyzed by GAP-PCR,hybridization and dideoxy chain termination method.Results The HbA1c values in non-hemoglobinophthy samples obtained using V Ⅱ-T 2.0,Ultra2,C2FP and PPI systems were well correlated with that of VⅡ system.The hemoglobin genotypes of the two cases of compand heterozygotes were aa/aa,βIVS-2-654/βNewYork and aa/aa,β41-42/βNewYork,respectively.The proportions of HbA were all 0 while the proportions of Hb NewYork were 93.5％ and 94.0％ respectively.The HbA1c results of the two cases detected were 4.3％ (23 mmol/mol)and 4.5％ (26 mmol/mol)by V Ⅱ,4.5 ％ (26 mmol/mol) and 4.6％ (27 mmol/mol) by VⅡ-T 2.0,no values and no values by C2FP,4.1％ (21 mmol/mol) and 4.3 ％ (23 mmol/mol) by Ultra2 and 4.2％ (22 mmol/mol) and 4.8 ％ (29 mmol/mol) by PPI systems,respectively.All the systems did not send warning for abnormal signs in profiles and results.Conclusion The five systems presented different early-warning ability for the double heterozygotes of Hb NewYork and β-thalassemia.The compand heterozygotes should theoretically not contain HbA1 c,but the systems V Ⅱ,V Ⅱ-T,Ultra2 and PPI showed detectable results of HbA1 c indicating all the four systems were not able to provide with early-warning.C2FP system did not report HbA1c result so it exhibited early-warning ability for the double heterozygote.

OBJECTIVETo investigate collagen type I alpha 1 (COL1A1) and alpha 2 (COL1A2) gene polymorphisms in Chinese and their relationship with bone mineral density.

METHODSTotalling 628 residents of Han nationality in Guangzhou aged 53.4-15.9 (range 20-79) years were surveyed for COL1A1 and COL1A2 gene genotypes by polymerase chain reaction-restriction fragment length polymorphism. Bone mineral density of the lumbar vertebrae, greater trochanter, femur neck and Ward's triangle was measured by dual-energy X-ray absorptiometry.

RESULTSCOL1A1 Sp1 polymorphism was not found in these subjects, and the genotype of all samples were type SS. COL1A2 genotyping revealed the distribution of EE genotype in 49.7%, Ee in 40.9% and ee in 9.4% of the subjects. The frequency distribution of EcoR1 alleles followed the Hardy-Weinberg equilibrium. The mean bone mineral density did no significantly differ among these genotype groups (P>0.05 by analysis of variance).

CONCLUSIONCOL1A1 Sp1 binding site polymorphism is absent and COL1A2 EcoR1 site polymorphism is not associated with bone mineral density in Chinese of Han nationality.

Adult ; Aged ; Bone Density ; China ; Collagen ; genetics ; Collagen Type I ; genetics ; Female ; Gene Frequency ; Genotype ; Humans ; Lumbar Vertebrae ; metabolism ; Male ; Middle Aged ; Polymorphism, Genetic ; Polymorphism, Restriction Fragment Length

Objective To establish a height prediction model of Chinese Han male based on the reported 547 height-associated single nucleotide polymorphisms (SNPs) loci in Europeans, and assess its accuracy for height estimation. Methods The DNA typing was analyzed in 59 Han male samples of Shandong province by Affymetrix SNP Array 6.0 chip and HiSeq 4000 sequencing platform. Prediction model was established using 547 height-associated SNPs loci as predictors and weight allele sums (WAS) as com-puting method. The accuracy of height prediction model was analysed using receiver operating characteris-tic (ROC) curve and area under curve (AUC). Results There was no height-associated SNPs locus was found by genome-wide association studies. In present study, height prediction model was established by WAS and obtained an AUC of 0.67 (95％CI:0.53-0.90). Conclusion It has reference value for predicting the height of Han male in Shandong province by WAS model based on 547 SNPs loci, while it is still necessary to further promote the accuracy of the prediction model by screening more height-associated SNPs loci with population heterogeneity.

OBJECTIVE: To determine the changes of mossy fiber sprouting in hippocampus of pre-kindling and post-kindling rats of chronic epilepsy induced by pentylenetetrazole (PTZ). METHODS: Sixty rats were randomly divided into a control group and a PTZ group (PTZ 30 mg/kg, intraperitoneal injection, once daily). The changes of mossy fiber sprouting in hippocampus of pre-kindling and post-kindling rats were examined by Timm staining. RESULTS: Before the occurrence of convulsion confirmed by behavior and EEG, the mossy fiber sprouting was found in the PTZ group. The grade of the mossy fiber sprouting increased with the gradual establishment of kindling effect. CONCLUSION Mossy fiber sprouting may play an important role in the onset and development of epilepsy.
Animals ; Disease Models, Animal ; Epilepsy ; chemically induced ; pathology ; Hippocampus ; drug effects ; pathology ; Kindling, Neurologic ; drug effects ; Male ; Mossy Fibers, Hippocampal ; growth & development ; Neurons ; metabolism ; Pentylenetetrazole ; pharmacology ; Rats ; Rats, Sprague-Dawley

OBJECTIVETo assess the value of fluorescence in situ hybridization (FISH) and bacterial artificial chromosome FISH (BAC-FISH) for the diagnosis for patients with marker chromosomes.

METHODSSixteen patients with marker chromosomes were analyzed with technologies including GTG-banding, Q-banding, multiplex FISH and BAC-FISH.

RESULTSThe marker chromosomes in the 16 patients were verified as der(Y) (2 cases), psu dic(Y) (1 case), psu dic(15) (1 case), dic(15) (1 case), del(Y) (1 case), r(X) (5 cases), i(14 or 22) (2 cases), i(18) (1 case).

CONCLUSIONFISH and BAC-FISH can both verify the origin of marker chromosomes and provide accurate information for the diagnosis and treatment of patients.

Adolescent ; Adult ; Child ; Chromosome Aberrations ; Female ; Genetic Diseases, Inborn ; diagnosis ; genetics ; Genetic Markers ; genetics ; Humans ; In Situ Hybridization, Fluorescence ; methods ; Male ; Young Adult