OBJECTIVETo clone the full-length Rcet3 gene, a novel gene related to family 2 cystatins, from mouse testis or other tissues.

METHODSRcet3 gene was cloned using digital differential display (DDD) and RT-PCR was performed for cloning the full-length Rcet3 gene from adult mouse testis cDNA library with sequence analysis.

RESULTSRcet3 cDNA was 610 bp in length, consisting of 4 exons to encode a protein with 140 amino acid residues. The encoded protein contained a potential signal peptide and a cystatin domain, but lacked critical consensus site important for cysteine protease inhibition. These characteristics could be seen in the Cres subgroup related to the family 2 cystatins. Rcet3 was specifically expressed in adult mouse testis, epididymis and the cerebrum, but at higher levels in the testis than in the epididymis and cerebrum.

CONCLUSIONRcet3 may be a new member of Cres subgroup of family 2 cystatins.

Amino Acid Sequence ; Animals ; Base Sequence ; Cloning, Molecular ; Cystatins ; genetics ; DNA, Complementary ; chemistry ; genetics ; Gene Expression Profiling ; Male ; Mice ; Mice, Inbred C57BL ; Molecular Sequence Data ; Reverse Transcriptase Polymerase Chain Reaction ; Sequence Analysis, DNA ; Testis ; metabolism

OBJECTIVETo establish a rabbit model of abdominal compartment syndrome (ACS) induced by prolonged intra-abdominal hypertension (IAH) and evaluate the therapeutic effect of somatostatin on ACS.

METHODSTwelve New Zealand rabbits were randomized equally into normal saline (NS) group and somatostatin group. ACS model was established by intra-abdominal bleeding (IAB) and intra-abdominal infusion with nitrogen gas to achieve an intra-abdominal pressure of 15 mmHg. The hemodynamics (SP, HR, CVP), hepatic function (ALT), renal function (BUN), antioxidation level (SOD, MDA) and blood electrolyte level (pH, [Na(+)], [Cl(-)], [CaNa(2+)], [KNa(+)]) of the rabbits were recorded 1-6 h after establishment of IAH.

RESULTSProlonged IAH caused decreased hemodynamic functions and antioxidation level as well as hyperkalemia and hypocalcemia (P<0.05), but these changes showed no significant differences between NS group and somatostatin group.

CONCLUSIONProlonged IAH causes cardiovascular function damages in rabbits possibly related to acidosis, electrolyte disturbances, and oxidative damage due to tissue ischemia and hypoxia. Somatostatin produces no obvious protective effects against the occurrence and progression of ACS.

Animals ; Disease Models, Animal ; Female ; Intra-Abdominal Hypertension ; drug therapy ; etiology ; physiopathology ; Male ; Rabbits ; Somatostatin ; therapeutic use

OBJECTIVETo research the bacteriostatic effects of Qingkailing Injection Extract (QKLIE) and combination therapy of Qingkailing Injection (QKLI) and antibiotics on bacteria carrying New Delhi metallo-3-lactamase 1 (NDM-1) blaNDM-1 resistance gene, and to determine their minimal inhibitory concentrations (MIC).

METHODSThe antimicrobial experiments of QKLIE (Radix Isatidis, baicalin, gardenia, honeysuckle) and combination therapy of QKLI and antibiotics were performed by using the agar dilution method and K-B method. The MIC was determined from each extract.

RESULTSThere were different degrees of inhibitory effects on resistant bacteria carrying blaNDM-1 by extracts from main components of QKLI. Of them, the inhibitory effect of baicalin was the best and the MIC of the resistant bacteria was 0.015 g/mL to WD, 0.020 g/mL to WX, 0. 005 g/mL to WJ, and more than 0.020 g/mL to pGEX-4T-NDM-1/DH5alpha (GST-NDM-1), respectively. The MIC value of each extract was sequenced from high to low as baicalin, honeysuckle, gardenia, and Radix Isatidis. Furthermore, combination therapy of QKLI and antibiotics greatly enhanced the antimicrobial activity of each antibiotics when used alone, showing very obvious antibacterial effects on multidrug resistant bacteria carrying blaNDM-1 gene. Of them, the optimal effects were obtained when combined with penicillins (penicillin G, mezlocillin, piperacillin/ tazobactam, ampicillin/sulbactam), with the antibacterial effects improved by 10 folds. The antibacterial effects of other kinds of antibiotics were improved to some extent. Conclusions QKLIE and combination therapy of QKLI and antibiotics showed better bacteriostatic effects on resistant bacteria carrying blaNDM-1 gene. This study provided theoretical bases for drug development, medication and treatment for super-resistant bacteria carrying blaNDM-1.

Anti-Bacterial Agents ; pharmacology ; Bacteria ; drug effects ; genetics ; Drug Resistance, Multiple, Bacterial ; drug effects ; genetics ; Drug Therapy, Combination ; Drugs, Chinese Herbal ; pharmacology ; beta-Lactamases ; genetics

OBJECTIVETo establish a rabbit model of abdominal compartment syndrome (ACS) and evaluate the impact of ACS on cardiovascular and respiratory functions and blood electrolyte levels in rabbits.

METHODSTwenty-four New Zealand rabbits were randomly allocated into 4 equal groups, namely the normal control group, ACS(5>\) group [intra-abdominal pressure (IAP)=5 mmHg], ACS(10) group (IAP=10 mmHg) and ACS(20) group (IAP=20 mmHg). ACS model was established by intra- abdominal bleeding (IAB) with intra-abdominal hypertension (IAH). All the data were recorded 1 h after inducing IAH including cardiovascular parameters (LVSP, LVEDP, ∓dp/dt max, SP, DP, HR, CVP), respiratory function (RR, PaO(2), PaCO(2), [HCO(3)(-)]), blood pH, and electrolyte level ([K(+)]).

RESULTSCompared with those in the normal control group, ACS20 group showed significantly decreased LVSP, LVEDP, ∓dp/dt max, SP, DP, HR, RR, PaO(2), [HCO(3)(-)], and blood pH but increased CVP, PaCO(2), and K(+) (P<0.05). In ACS(10) group, all the parameters except for RR and PaO(2) showed similar changes as seen in ACS(20) group (P<0.05) but with lower amplitudes of variations. In ACS(5) group, only LVSP and HR were reduced remarkably (P<0.05) while the other parameters showed no significant variations.

CONCLUSIONIAB plus IAH may cause damage to the cardiovascular and respiratory functions and lead to ACS in rabbits.

Abdominal Cavity ; physiopathology ; Animals ; Blood Gas Analysis ; Disease Models, Animal ; Intra-Abdominal Hypertension ; blood ; physiopathology ; Rabbits ; Respiratory System ; physiopathology ; Ventricular Function, Left

OBJECTIVETo investigate the relationship between wedge-shaped defects and occlusal interference.

METHODSFollowing examination from 46 patients, a total of 157 teeth were identified to have the criteria set for wedge-shaped defects and regarded as the experiment group. Also, 157 adjacent teeth exhibiting no such noncarious cervical lesions were randomly selected from the same group of patients and regarded as the control group. The distribution of occlusal force and time were examined with T-Scan II system in 46 patients with wedge-shaped defects in intercuspal occlusion position, protrusive movement and lateral movement. Occlusal interference and premature contact were evaluated and compared between the two groups.

RESULTSThe proportion of the teeth with premature contact in experiment group was 6.37%, while the control group was 2.55%, there was no significant difference between the two groups (P > 0.05). The total proportion with occlusal interferences in experiment group was 23.57%, which was significantly higher than that of the control group (10.19%, P < 0.05), in experiment group the proportion with working side interferences was 15.92%, and in control group, the proportion was 3.82%, there was significant difference between the two groups (P < 0.01). At the same time, the teeth with occlusal interferences had more serious degree of the wedge-shape defects than those with no occlusal interference.

CONCLUSIONThe increased occlusal force has relation to the formation as well as severity of wedge-shaped defects.

Bicuspid ; Bite Force ; Dental Occlusion ; Humans ; Male ; Tooth Abrasion

Objecfive To establish a method for in vitro culture and identification of neural stem cells(NSCs)derived from the olfactory bulb(OB)of adult mice and test the possibility of the OB as a new source of seed cells of adult NSCs. Methads NSCs were isolated from the OB of adult mice and cultured in serum-free medium.Clonal culture and BrdU incorporation assay were performed to assess the self-renewal and proliferative activities of the NSCs.Fluorescence immunocytochemistry was carried out to examine the expression of the NSC markers nestin and SOX2,neuronal marker Tujl,astrocyte marker GFAP and oligodendroeyte marker 04. Results NSCs possessing self-renewal and proliferative capacities were obtained from the OB of adult mice,and the cells grew in the form of floating neurospheres in the medium.The neurospheres consisted of cells were positive for NSC markers nestin and SOX2,which Were able to differentiate into Tuj1-positive neurons,GFAP-positive astrocytes and 04-positive oligodendrocytes. Conclusion NSCs are present in the OB of adult mice,and the NSCs isolated from the OB can proliferate and differentiate in vitro with obvious stem cell properties.suggesting the feasibility of using OB as anew source of adult NSCs.

FMS-like tyrosine kinase 3 (FLT3) is a receptor of tyrosine kinase that is constitutively activated in most of acute myeloid leukemia patients and seems to give an adverse prognosis. In order to explore the silencing effect of FLT3 targeted short hairpin RNA (FLT3-shRNA) on acute leukaemia cell line THP-1, three FLT3-shRNAs (shRNA1, shRNA2, shRNA3) were designed and synthesized by transcription system in vitro and then transfected into THP-1 cells. FLT3 mRNA was analyzed by semi-quantitative RT-PCR, FLT3 protein was detected by Flow cytometry and immunofluorescence. The results indicated that FLT3 expression was downregulated by shRNA1 and shRNA3, and shRNA1 showed stronger inhibitory effect. At 48 hours following transfection, the inhibitory rate of 25 nmol/L shRNA1 was 72.95 +/- 2.07%, lasting 72 hours. The 5 nmol/L and more concentration of FLT3 shRNA1 could downregulate FLT3 mRNA level, which displayed a quantity-effect relation; the inhibitory rate of 15 nmol/L shRNA1 was 67.53 +/- 0.66%. FLT3 protein was located on THP-1 cell membrance, its expression was downregulated obviously by shRNA1, at 72 hours following transfection the inhibitory rate of shRNA1 was 79.67 +/- 0.66%. shRNA1 showed the best inhibitory effect on FLT3 protein, the optimal time of which was 72 hours with an inhibitory rate of 79.67%. It is concluded that FLT3-shRNA1 shows a desireable FLT3-targeted inhibitory effect, which can be used for further investigation of FLT3 mechanism or FLT3 targeting treatment.
Humans ; Leukemia, Myeloid, Acute ; genetics ; metabolism ; RNA Interference ; RNA, Messenger ; genetics ; RNA, Small Interfering ; genetics ; Transcription, Genetic ; Tumor Cells, Cultured ; fms-Like Tyrosine Kinase 3 ; genetics

OBJECTIVETo investigate foot biomechanics characteristic of patients with type 2 diabetes mellitus.

METHODSThis study was conducted among 303 patients with type 2 diabetes. The whole foot was divided into 10 regions, namely the first toe (T1); the second to fifth toes (T2-5); the first, second, third, fourth, and fifth metatarsals (M1, M2, M3, M4, and M5, respectively); midfoot (MF), and the heel medial (HM). Foot arch index, foot angle and maximum peak pressure (MPP) of the 10 regions were measured using a Footscan gait system.

RESULTSThe maximum peak pressure of 10 regions decreased in the order of M3>M2>HM>M4>HL>M1>M5>T1>ML>T2-5 for the left foot, and in the order of M3>M2>HM>M4>HL>M1>M5>T1>ML>T2-5 for the right foot. The MPP in M1 region was higher in the right than in the left foot (P<0.05). The MPP in M3, M4, M5, and MF was higher in the left than in the right foot (P<0.05). The percentage of high-risk foot (defined by a total plantar pressure ≥70 N/cm) was 34% on the left and 17.7% on the right. An increased BMI was associated with a significant increase in high-risk foot, but not for the right foot in underweight patients. Foot flat phase was extended and forefoot push-off phase shortened in stance phase in the patients. Compared with the right foot, the left foot showed a significantly increased foot arch index and increased low and high arch rates with a decreased normal arch rate. Total plantar pressure was higher in of the left high arch foot than in normal arch foot. The foot angle was significantly larger on the right than on the left. The bilateral total plantar pressures were significantly greater in male patients (P<0.05) and increased with age but were not associated with the duration of DM, foot angle, or glycosylated hemoglobin level.

CONCLUSIONDiabetic patients have obvious alterations in foot biomechanics with abnormalities of the plantar pressure, and the percentage of high-risk foot increases in overweight and obese patients, suggesting the need of body weight control in these patients when administering offloading treatment for prevention of diabetic foot ulcer.

Biomechanical Phenomena ; Diabetes Mellitus, Type 2 ; physiopathology ; Diabetic Foot ; prevention & control ; Female ; Foot ; physiopathology ; Gait ; Heel ; physiopathology ; Humans ; Male ; Obesity ; physiopathology ; Overweight ; physiopathology ; Pressure

OBJECTIVEFMS-like tyrosine kinase 3 (FLT3) is a receptor tyrosine kinase that is constitutively activated in (70-90)% pediatric patients with acute myeloid leukemia (AML) and appears to confer an adverse prognosis. Although several FLT3-selective small molecule inhibitors and antibodies were developed with varied degrees of success, to address the specificity and resistance, new approaches for specifically targeted FLT3 are needed and RNA interference is a promising choice. The aim of the present study was to investigate the efficacy of suppression of FLT3 induced by small hairpin interfering RNA (shRNA) on myeloproliferation and apoptosis in an acute monocytic leukemia (AMOL) cell line THP-1.

METHODSFLT3-targeted small hairpin interfering RNA (FLT3-shRNA) was designed and synthesized by transcription system in vitro was transfected into THP-1 cells. Firstly FLT3 mRNA level was detected by semi-quantitative RT-PCR and FLT3 protein level was detected by flow cytometry (FCM) to verify the efficacy on FLT3-shRNA interference at 48 h after transfection. Cell growth viability was measured at 24 h, 48 h and 72 h after treatment with CCK-8. The distribution of cell cycle was assayed by FCM, and apoptosis was analyzed by DNA Ladder and Annexin V-FITC Staining at 48 h.

RESULTSFLT3 targeted shRNAs was synthesized successfully and the concentration of 15 nmol/L for 48 h could obtain desirable downregulation of FLT3 expression, the inhibitory percentages of FLT3 mRNA and protein were (72.95 +/- 2.07)% and (65.39 +/- 5.57)%, respectively. The suppression of FLT3 induced by FLT3-shRNA resulted in marked inhibition of cell growth and the inhibitory percentages were (36.66 +/- 3.67)% at 48 h, (35.56 +/- 0.73)% at 72 h. FLT3-shRNA induced the inhibition of cell cycle from G(0)/G(1) phase to S phase, the percentage of sub-G(0)/G(1) phase (65.71 +/- 4.47)% was higher than those in the PBS-control group (52.23 +/- 2.98)%, NC-shRNA control group (51.81 +/- 1.44)%, P < 0.01; the percentage of S phase (25.11 +/- 2.70)% was lower than those in the PBS-control group (34.41 +/- 4.07)% and NC-shRNA control group (32.50 +/- 1.46)%, P < 0.05. Furthermore treatment with FLT3-shRNA for 48 h resulted in clear apoptosis ladder, the percentage of early apoptosis detected by Annexin V-FITC was (18.59 +/- 2.07)% which was significantly higher than that in the PBS-control group (4.00 +/- 0.50)% and the NC-shRNA control group (6.06 +/- 0.70)%, P < 0.001.

CONCLUSIONThe suppression of FLT3 induced by the shRNA can effectively inhibit cell proliferation, and apoptosis induction on THP-1 cells, which indicates that this approach may bear the therapeutic potential on childhood AMOL.

Apoptosis ; drug effects ; genetics ; Cell Proliferation ; drug effects ; Child ; Humans ; Leukemia, Monocytic, Acute ; enzymology ; pathology ; Protein-Tyrosine Kinases ; metabolism ; RNA Interference ; physiology ; RNA, Small Interfering ; pharmacology ; Receptor Protein-Tyrosine Kinases ; metabolism ; fms-Like Tyrosine Kinase 3 ; metabolism

BACKGROUNDIncreasing prevalence of Staphylococcus aureus (S. aureus), particularly methicillin-resistant S. aureus (MRSA) has been reported in China. In this study, we investigated the drug resistance characteristic, genetic background, and molecular epidemiological characteristic of S. aureus in Changsha.

METHODSBetween January 2006 and December 2008, 293 clinical isolates of S. aureus were collected from 11 hospitals in Changsha and identified by the Vitek-2 system. All the isolates were verified as MRSA by PCR amplification of both femA and mecA genes. K-B disk method was used to test drug sensitivity of S. aureus to antibiotics. Pulsed-field gel electrophoresis (PFGE) was performed for genotypic and homologous analysis of 115 isolates randomly selected from the original 293 clinical S. aureus isolates.

RESULTSS. aureus was highly resistant to penicillin, ampicillin, erythromycin, and clindamycin with resistant rates of 96.6%, 96.6%, 77.1%, and 67.2% respectively. All the isolates were susceptible to tecoplanin, vancomycin, and linezolid. MRSA accounted for 64.8% (190/293) of all the S. aureus strains. The 115 S. aureus isolates were clustered into 39 PFGE types by PFGE typing, with 13 predominant patterns (designated types A to M) accounting for 89 isolates. The most prevalent PFGE type was type A (n = 56, 48.7%) and 100.0% of type A strains were MRSA. PFGE type A included 13 subtypes, and the most prevalent subtype was subtype A1 (46.4%, 26/56). Strains with PFGE type A were isolated from eight hospitals (8/11), and both subtypes A1 and A4 strains were isolated in a university hospital.

CONCLUSIONSClinical isolates of S. aureus in Changsha were resistant to multiple traditional antibiotics. There was an outbreak of PFGE type A MRSA in this area and the A1 subtype was the predominant epidemic clone. Dissemination of the same clone was an important reason for the wide spread of MRSA.

Ampicillin ; pharmacology ; Anti-Bacterial Agents ; pharmacology ; China ; Clindamycin ; pharmacology ; Electrophoresis, Gel, Pulsed-Field ; Erythromycin ; pharmacology ; Humans ; Methicillin-Resistant Staphylococcus aureus ; drug effects ; genetics ; Microbial Sensitivity Tests ; Penicillins ; pharmacology ; Staphylococcus aureus ; drug effects ; genetics ; Vancomycin ; metabolism