Objective To observe the effects of enzymolysis of human B like antigen by ? galactosidase treatment on the morphology and function of cynomolgus monkey red blood cell (RBC) and to evaluate the safety of the enzyme treated RBC in animal transfusion. Methods RBC with human B like antigen was treated by recombined ? galactosidase and the effect was evaluated by absorption elution test. Meanwhile, morphology and function of the treated RBC were examined and compared with pre treatment RBC. ? galactosidase treated RBC was labeled with FITC and then infused to blood group A cynomolgus monkeys. Flow cytometry was used to detect the survival of donor RBC in recipient’s blood. Blood chemistry and urinalysis of recipients were performed before and after transfusion. Results The human B like antigen of cynomolgus monkey RBC was obliterated by ? galactosidase treatment and the treated RBC maintained normal morphology and function. Survival at 24 h after transfusion was 84.6% and 68.1%. T 1/2 were 7d and 8d versus 13d in the controls. There were no change in blood chemistry and urinalysis of the recipients. Conclusion Enzymolysis of cynomolgus monkey RBC does not affect the cellular function and morphology. Transfusion of the enzyme treated human B like RBC into human A like cynomolgus monkeys is safe.

OBJECTIVETo explore the effects of microtubule depolymerization (MD) on the spontaneous beating rate, action potential (AP), and oxygen consumption of cardiac myocytes in rats and its mechanism.

METHODSOne-hundred and eighty neonatal SD rats divided into 12 batches were used in the experiment, and 15 rats in each batch were sacrificed for the isolation and culture of cardiac myocytes after the heart tissues were harvested. The cardiac myocytes were respectively inoculated in one 12-well plate filled with 6 round cover slips, one 12-well plate filled with 6 square cover slips, two cell culture flasks, and two cell culture dishes. After routine culture for three days, the cardiac myocytes from all the containers were divided into normal control group (NC, routinely cultured with 3 mL DMEM/F12 solution rewarmed at 37 °C for 3 h) and group MD (routinely cultured with 3 mL DMEM/F12 solution rewarmed at 37 ° and containing 8 µmol/L colchicine for 3 h) according to the random number table, with 3 holes, 1 flask, or 1 dish in each group. The morphological changes in microtubules were observed with confocal laser scanning microscope after immunofluorescent staining. The content of polymerized or dissociative α-tubulin was determined by Western blotting. Spontaneous beating rate of the cells was observed and calculated under inverted microscope. Dissolved oxygen concentration of DMEM/F12 solution containing cardiac myocytes was determined by oxygen microelectrode system before and after the addition of colchicine. Additionally, dissolved oxygen concentration of DMEM/F12 solution and colchicine + DMEM/F12 solution was determined. The whole-cell patch-clamp technique was used to record AP, delayed rectifier K+ current (I(K)), and L-type Ca2+ current (I(Ca-L)) in cardiac myocytes; current density-voltage (I-V) curves were drawn based on the traces. Data were processed with independent or paired samples t-test.

RESULTS(1) In group NC, microtubules of cardiac myocytes were around the nucleus in radial distribution with intact and clear linear tubiform structure. The microtubules in group MD were observed in dispersive distribution with damaged structure and rough linear tubiform structure. (2) In group MD, the content of dissociative α-tubulin of cells (0.61 ± 0.03) was obviously higher than that in group NC (0.46 ± 0.03, t = -6.99, P < 0.05), while the content of polymerized α-tubulin (0.57 ± 0.04) was significantly lower than that in group NC (0.88 ± 0.04, t = 9.09, P < 0.05). (3) Spontaneous beating rate of cells was (59 ± 8) times per min in group MD, which was distinctly higher than that in group NC [(41 ± 7) times per min, t = 5.62, P < 0.01]. (4) Dissolved oxygen concentration of DMEM/F12 solution containing cardiac myocytes was (138.4 ± 2.5) µmol/L, and it was reduced to (121.7 ± 3.6) µmol/L after the addition of colchicine ( t = 26.31, P < 0.05). There was no obvious difference in dissolved oxygen concentration between DMEM/F12 solution and colchicine + DMEM/F12 solution (t = 0.72, P > 0.05). (5) Compared with that of group NC, AP morphology of cells in group MD changed significantly, with unobvious repolarization plateau phase and shorter action potential duration (APD). The APD20, APD50, and APD90 were respectively (36.2 ± 3.8), (73.7 ± 5.7), and (115.1 ± 8.0) ms in group MD, which were significantly shorter than those of group NC [(40.2 ± 2.3), (121.4 ± 7.0), and (169.4 ± 5.6) ms, with t values respectively 2.61, 15.88, and 16.75, P values below 0.05]. (6) Compared with that of group NC, the I-V curve of I(K) of cells in group MD moved up with higher current density under each test voltage (0 to 40 mV) after activation ( with t values from 2. 70 to 3. 76, P values below 0.05) . (7) There was not much alteration in current density of I(Ca-L) under each test voltage (-30 to 50 mV) between 2 groups (with t values from -1.57 to 1.66, P values above 0.05), and their I-V curves were nearly overlapped.

CONCLUSIONSAfter MD, the I(K) is enhanced without obvious change in I(Ca-L), making AP repolarization faster and APD shortened. Then the rapid spontaneous beating rate increases oxygen consumption of cardiac myocytes of rats.

Action Potentials ; Animals ; Cells, Cultured ; Energy Metabolism ; Microtubules ; metabolism ; Mitochondria, Heart ; metabolism ; Myocytes, Cardiac ; metabolism ; Rats ; Rats, Sprague-Dawley ; Tubulin ; metabolism

AIM: To study the effect of insulin on proliferation and hypertrophy of cardiac myocytes and its role in the induction of cardiac hypertrophy. METHODS: 1. The neonatal rat cardiac myocytes and cardiac fibroblasts were cultured respectively and identified with light microscopy, electron microscopy and immunocytochemistry. 2. Cell proliferation was measured with cell number, metabolic activity and DNA synthesis (with WST-1, BrdU enzyme-linked immunosorbent assay ) and the percentage of S+G 2+M in cell cycle (by flow cytometry ). 3.Cell hypertrophy was evaluated by cell protein content (Coomassie Briliant Blue's method). RESULTS: 1. The cultured cells showed the characteristic of cardiac myocytes and cardiac fibroblasts, respectively. 2. After being treated with insulin, the cell number, absorbance of BrdU incorporation and WST-1 cleavage products and the percentage of S+G 2+M of cardiac fibroblasts increased significantly ( P 0.05). 3. Protein content of cardiac myocytes increased significantly in a dose-dependent manner ( P

Objective To analyze epidemiological characteristics of mitochondrial DNA12SrRNA A1555G mutation in Chinese populations with non-syndromic sensorineural hearing loss by the literature review and find the main actual deficiencies in course of epidemiological study.Methods From Cbmdisc and PUBMED database pulled out were all published epidemiological literatures about Chinese mtDNA12SrRNA A1555G mutation from 1996 to 2008.Reviewed were the primary data of these studies including the number of samples,demographic characteristics of the samples,mutation frequencies,interrelations between the mutation and aminoglycoside exposure and so on.Results 21 papers out of 25 were induded in this study.The patients had non-syndromic sensorineural hearing loss from 14 regions of China.A total of 3 473 were found including 230 patients with A1555G mutation and the average mutation frequency was 6.62%.The samples in each regions ranged from 72 to 802 and the reported mutation frequencies were from 0.67%-14.6%.The statistical discrepancy was significant among mutation frequencies in different regions by χ~2 test(P=0.0000).The number of patients with aminoglycoside antibiotics exposure was 739 including 100 with A1555G mutation in all literatures.The proportions in different regions were from 2.70% to 33.33% with the average of 13.53%.The average proportion was significantly higher than the mutation frequency in patients with non-syndromic sensorineural hearing loss.Conclusion Some deficiencies in epidemiological research Omutation in China included age,ethnic,and geographic bias,insufficiency of samples,inadequate randomization and so on.Researchers should focus with more efforts on the epidemiological characteristics of A1555G mutation in Chinese people.

OBJECTIVE To investigate the contribution of the GJB6 gene [encoding connexin 30 (C?30)] mutations in Chinese population with sporadic non-syndromic hearing impairment. METHODS PCR reactions were performed with two pair of primers for the coding sequence of GJB6 gene and for the deletion of GJB6. PCR products bidirectional sequencing was subsequently applied in 214 patients with hearing loss and 86 normal controls. RESULTS A novel heterozygous mutation-233(C→A) was found, which results in amino acid change, A78D. This mutation wasn't detected in the control subjects. The altered valine residue lies within the second conserved transmembrane domain. The large deletion△(GJB6/ D13S1830)] of GJB6 was not found in this group. CONCLUSION The large deletion of GJB6 was not found in the Chinese deafness population. A novel heterozygous mutation of GJB6 was found. These results indicated GJB6 mutations are not a major cause of hearing loss in the Chinese population.

Objective To establish a in vitro cell model of inflammation following ischemic injury in rat astrocytes, and study the effects of mild hypothermia on the response of astrocytes to ischemia and inflammation. Methods Primary cultured neonatal SD rat astrocytes identified by GFAP immunofluorescence staining were divided into normal control group (C), hypoxia group (H), and hypoxia and leukocyte treatment group (H+W). Hypoxic injury of the cells was induced by exposure to glucose-flee DMEM medium in 5% CO2 and 95% N2 for 4 h, and for leukocyte treatment, the astrocytes were co-cultured with leukocytes isolated from rat blood. The 3 groups of cells were subjected to hypothermia treatments at 37, 34, 32 and 30 ℃, and the cell viability was estimated by lactic dehydrogenase (LDH) release assay and live/dead assay. Results Ischemic injury occurred in the astrocytes after 4 h of hypoxic exposure. Hypoxic treatment of the cells at 37 ℃ resulted in increased LDH release in all the 3 groups, and the increment intensified in the order of groups C, H and H+W. Mild hypothermia at 34 ℃ and 32 ℃ significantly reduced the LDH release in groups H and H+W (P<0.05), but hypothermia at 30 ℃ significantly increased LDH release in comparison with that at 32 ℃ (P<0.05). Conclusion Mild hypothermia can significantly reduce the severity of ischemic and inflammatory injuries in rat astrocytes possibly mediated also by other mechanisms in addition to inhibited astrocyte metabolism due to low temperatures.

AIM　To study the apoptosis and characterizati on of cultured vascular smooth muscle cells (VSMCs) induced by cholestan-3β,5 α,6β-triol(Triol) and compared with 25-hydryoxycholestrol(25-OH). M ETHODS　The culture of vascular muscle smooth cells (VSMCs), light and e lectron transmission microscopy and TdT-mediated dUPT nick-end labeling (TUNEL ) technique. RESULTS　After being treated with oxyterols, VSMCs showed apoptosis of ultrastracture change including shrinkage, condensation of nuclear chromatin, fragmentation nuclei and formation of apoptotic body. TUNEL r evealed that Triol-induced apoptosis in VSMCs was in a concentration-dependent manner. The effect of Triol and 25-OH at a 30 μmol*L-1 concentration i n culture medium induced apoptosis of VSMCs, the former was not but the latter was inhibited by cholesterol at a 50 μmol*L-1 concentration. CO NCLUSION Triol can induce VSMCs apoptosis in vitro and oxysterol-i nduced apoptosis in VSMCs may be mediated through various pathway and different mechanism. Oxysterol-induced apoptosis in VSMCs may play an important role in t riggering atherosclerosis plaque rupture and result to the onset of the acute co ronary syndromes.

To study the chemo-preventive effect of the supercritical extracts from Angelica sinensis (SFE-AS) on induced colorectal carcinoma in mice by using the AOM/DSS-induced male mice colorectal carcinoma model, and discuss its possible action mechanism. Male Balb/c mice were subcutaneously injected with single dose of azoxymethane (AOM, 10 mg x kg(-1) body weight). One week later, they were given 2% dextran sodium sulfate (DSS) in drinking water for 7 days to induce colorectal carcinoma. Each drug group was orally administered with supercritical extracts from Angelica sinensis at 15, 30, 60 mg x kg(-1) until the 17th week. The tumor incidence rate of the SFE-AS group, mice tumor-bearing quantity and tumor-bearing volume of the SFE-AS group were lower than that of the AOM/DSS model control group, which may be related with the significant reduction of PCNA, COX-2, iNOS in the AOM/DSS-induced mouse colorectal carcinoma model associated with inflammation by SFE-AS. According to the results of this study, SFE-AS showed an intervention effect in the incidence and development of AOM/DSS-induced mouse colorectal carcinoma associated with inflammation, and could be further used in chemo-preventive studies on human colorectal carcinoma.
Angelica sinensis ; chemistry ; Animals ; Azoxymethane ; adverse effects ; Colonic Neoplasms ; chemically induced ; genetics ; immunology ; prevention & control ; Colorectal Neoplasms ; chemically induced ; genetics ; immunology ; prevention & control ; Cyclooxygenase 2 ; genetics ; metabolism ; Dextran Sulfate ; adverse effects ; Disease Models, Animal ; Drugs, Chinese Herbal ; administration & dosage ; Humans ; Male ; Mice ; Mice, Inbred BALB C ; Proliferating Cell Nuclear Antigen ; genetics ; immunology

OBJECTIVETo explore the role of Compound Danshen Injection (CDI) in regulating the expression of aquaporin 3 (AQP3) in human amnion epithelium cells (hAECs), and to study the relation between c-Jun N-terminal kinase (JNK) signal pathway and AQP3.

METHODShAECs were isolated and primarily cultured from term pregnancy with normal amniotic fluid volume and from term pregnancy with oligohydramnios, and then hAECs were further divided into four groups, i.e., the blank control group (A), the SP600125 group (B), the CDI group (C), and the SP600125 +CDI group (D). The cell viability was measured by cell counting kit-8 assay (CCK-8). The expression of total JNK, phosphorylated JNK, and AQP3 were determined by Western blot.

RESULTS(1) In hAECs with normal AFV or with oligohydramnios: There was no statistical difference in the cell viability or the expression of total JNK among the 4 groups (P > 0.05). But there was statistical difference in the expression of p-JNK (P < 0.05). Compared with A group, the expression of p-JNK was obviously down-regulated in B group, but obviously up-regulated in C group (P < 0.05). The expression of p-JNK was significantly lower in D group than in C group, but higher than that in A group or B group (P < 0.05).The AQP3 expression in the hAECs with normal amniotic fluid volume of C group and D group were higher than that in the A group (P < 0.05). However, there was no statistical difference in the AQP3 expression between C group and D group (P > 0.05). In hAECs with oligohydramnios, the expression of AQP3 obviously decreased in B group, but up-regulated in C group (both P < 0.05). The expression of AQP3 was lower in D group than in C group, but higher than in B group (P < 0.05).

CONCLUSIONCDI could regulate the AQP3 expression in hAECs with oligohydramnios via activating the JNK signal pathway.

Amnion ; cytology ; drug effects ; Aquaporin 3 ; metabolism ; Cells, Cultured ; Drugs, Chinese Herbal ; pharmacology ; Epithelial Cells ; drug effects ; metabolism ; Female ; Humans ; JNK Mitogen-Activated Protein Kinases ; metabolism ; MAP Kinase Signaling System ; physiology

Objective: To explore the correlation between malocclusion and body image issues in college students. Methods: A total of 1 851 students in three universities in Jingmen were selected by using stratified cluster sampling method. Angle s classification of malocclusion was used to determine the number of three types of malocclusions. Body image issues were self reported and its relationship with different types of malocclusions was explored. Results: The proportions of Classes Ⅰ, Ⅱ and Ⅲ malocclusion in college students with malocclusion were 71.21%, 16.32%, and 12.47%, respectively. The detection rates of body image issues among students with Classes Ⅰ,Ⅱ and III malocclusions were 36.64%, 54.78% and 65.83%, respectively. No significant difference were found in the detection rates of sexual organ issues and gender issues in college students with different types of malocclusions( χ 2= 0.75, 0.53, P >0.05). There were significant differences in the detection rates of appearance troubles (27.59%, 33.12%, 50.83% ) and stature troubles ( 24.09% , 31.21%, 44.17%) in students with Classes Ⅰ, Ⅱ and Ⅲ malocclusions( χ 2=5.62, 2.89, P <0.05). Conclusion The prevalence of body image issues in college students increases with severity of malocclusions. Appearance and stature troubles are issues mostly concerned among college students. Psychological evaluation for students with Class Ⅲ malocclusion should be especially emphasized when administrating orthodontic treatment.