[Observation on the efficacy of CT-guided lumbar sympathetic chemical destructive block in the treatment of cold sensation of limbs].

Objective: To observe the clinical effects of CT-guided chemical destructive block of lumbar sympathetic nerve in the treatment of cold sensation of limbs. Methods: In this retrospective analysis, clinical data of 43 patients with cold sensation of limbs treated by lumbar sympathetic chemical destructive block in the Affiliated Hospital of Jiaxing University from January 2015 to January 2018 were collected. The changes of heart rate, non-invasive blood pressure (NIBP), oxygen saturation (SpO(2)), plantar temperature and peripheral perfusion index (PI) of patients were recorded and analyzed before treatment and 5 min after injection of anhydrous ethanol. The patients were followed up at postoperative 1 day, 1 week, 1 month, 3 months, 6 months, 1 year and 2 years. Results: Fourty-three patients underwent bilateral lumbar sympathetic nerve chemical destructive block under the CT-guided, and all patients were punctured to the target successfully. The PI of patients before and after treatment were 1.2±0.6, 7.2±3.0 respectively, which was significantly increased after treatment compared with before treatment, and the difference was statistically significant (t=12.386, P<0.05). The plantar temperature of patients before and after treatment respectively were (29.6±1.7)℃, (34.6±1.1)℃, which was significantly increased after treatment compared with before treatment, and the difference was statistically significant (t=15.057, P<0.05). There were no significant differences in heart rate, NIBP and SpO(2) between before and after treatment (all P>0.05). Lumbar sympathetic chemical destructive block was clinically effective in 39 patients (90.7%) and ineffective in 4 patients (9.3%). Among the 39 clinically effective patients, the curative effects were excellent in 29 cases and improved in 10 cases. Postoperative recurrence occurred in 10 cases (25.6%). The satisfaction rates of patients at 1 day, 1 week, 1 month, 3 months, 6 months, 1 year and 2 years after operation were 93.0%, 90.7%, 86.0%, 76.7%, 69.7%, 65.1% and 53.4%, respectively. Conclusion: Lumbar sympathetic chemical destructive block is a safe and effective way for the treatment of cold sensation of limbs, which can improve the symptoms of cold sensation of limbs to some extent.

[Effects of lateral and medial points of thoracic paravertebral nerve block by ultrasound for rapid recovery after laparoscopic cholecystectomy].

Objective: To investigate the effects of ultrasound-guided lateral and medial point blocks of thoracic paravertebral space on the rapid recovery of laparoscopic cholecystectomy. Methods: A total of 90 patients of either sex, aged 18-67 years, weighted 45.10-91.80 kg, of American Society of Anesthesiologists physical status Ⅰ or Ⅱ, undergoing elective laparoscopic cholecystectomy were divided into two groups (n=45) using a random number table: lateral point group of thoracic paravertebral space (group A) and medial point group of thoracic paravertebral space (group B). Ultrasound-guided thoracic paravertebral nerve block was performed before induction of general anesthesia. The puncture point of group A was positioned as the intercostal block of the thoracic paravertebral space of the right side of T(6)-T(11), and the puncture point of the group B was positioned as the thoracic paravertebral body of the right side of T(6)-T(11) thoracic paravertebral space. The thoracic paravertebral block was performed with 2 ml of 0.75% ropivacaine per injection for a total of 10 ml. The visual analog scale (VAS) scores of resting pain and active pain at 4, 8, 12 and 24 h after operation were observed. The anus recovery time after surgery and perioperative hypotension were also recorded. Results: The blood pressures in group A were significantly higher than those in group B at 4, 8, 12 and 24 h after operation, which were(73±7) vs (70±7), (78±7) vs (74±7),(82±7) vs (79±7),and (87±7) vs (83±7) mmHg,and the differences were statistically significant (t=2.29, 2.54, 2.33, 2.37, all P<0.05). The VAS scores of resting pain and active pain in group A were significantly higher than those in group B, and the differences were statistically significant (Z=-2.29, -2.51, -2.21, -2.39, -2.53, -2.25, -2.30, -2.24, all P<0.05). The postoperative anal exhaust recovery time of the patients in group A was (21.8±1.9) min that was obvious lower than that in group B which was (22.7±1.9) min with statistically significant difference (t=2.12, P<0.05). There was no significant difference in the incidence of postoperative dizziness, nausea, vomiting, and pruritus (χ(2)=0.28, 0.72, 0.45, 0.21,all P>0.05). Conclusions: In the procedure of thoracic paravertebral block under the guidance of ultrasound, the closer blocking points are to the central axis of the spine, the better the postoperative analgesic effect, but the more obvious the postoperative blood pressure reduction and the longer the anal recovery exhaust time. The further away from the central axis of the spine, the more analgesic effect decreases, but the blood pressure decreases and the anal recovery time is relatively rapid.

[Study of genetic etiology in fetuses with severely short limbs in the first and second trimester using whole exome sequencing].

Objective: To investigate pathogenic genes related to the phenotype of fetus with severely short limbs in the first and second trimester by whole exome sequencing (WES). Methods: Thirteen fetuses with severely short limbs detected by ultrasonography in the first and second trimester admitted in Chinese PLA General Hospital from September 2016 to June 2018 were collected. All cases were performed induced abortion, 6 of which were carried out karyotype analysis of amniotic fluid at the same time. WES and copy number variations (CNV) were performed on specimens from fetal tissues after labor induction. The suspected pathogenic mutations were validated by Sanger sequencing reactions. Results: No abnormal karyotypes or pathological CNV were found. In 10 fetuses, pathogenic or possibly pathogenic mutations were detected in the following genes: COL2A1, FGFR3, COL1A1, COL1A2, DYNC2LI1 and TRIP11, all of which were essential to skeletal development. The diagnostic yield of WES in the fetuses with severe short limbs was 10/13. Conclusions: In the first and second trimester, most of the fetuses with extremely short limbs suffer from monogenic diseases. WES is likely to be a valuable diagnostic testing option for the fetuses with severe short limbs.

[Initial study of transthoracic echocardiography guided three-dimensional printing on the application of assessment of structural heart disease].

Objective: To investigate the feasibility and diagnostic value of preoperative transthoracic echocardiography guided three dimensional printing model (TTE Guided 3DPM) on the assessment of structural heart disease (SHD). Methods: From February 2016 to October 2016, 44 patients underwent cardiac surgery in Tianjin Chest Hospital, forty-four patients were assessed preoperatively using TTE Guided 3DPM, including 25 males and 19 females, aged 3-75 years, with an average of (44±22) years. compared to conventional three dimensional transthoracic echocardiography (3D-TTE), and took direct intraoperative findings as "Golden Standard" simultaneously. There are twelve patients with SHD, including four cases with mitral prolapse, two cases with partial endocardial cushion defect, two cases with secondary atrial septal defect, two cases with rheumatic mitral stenosis, one case with tetralogy of Fallot, one case with ventricular septal defect (VSD), thirty-two patients without SHD were designed as negative control. Results: The sensitivity and specificity of TTE Guided 3DPM were greater than or equal to 3D-TTE, P value of McNemar test of 3D-TTE was greater than 0.05, the difference was not statistically significant, kappa=0.745, P<0.01, indicated that the results of 3DTTE and the gold standard were generally consistent.P value of McNemar test of TTE Guided 3DPM was greater than 0.05, the difference was not statistically significant, kappa=0.955, P<0.01, indicated that the results of TTE Guided 3DPM and gold standards were consistent. Compared with 3D-TTE and TTE Guided 3DPM, P value was greater than 0.05, the difference was not statistically significant, kappa=0.879, P<0.01, indicated that the results of 3D-TTE and TTE Guided 3DPM were consistent. TTE Guided 3DPM displayed the three-dimensional structure of SHD cardiac lesions clearly, which were consistent with intraoperative findings. Conclusion: TTE Guided 3DPM provides essential information for the preoperative evaluation and decision of SHD.

[Clinical analysis of 21 cases with short fetal femur in the third trimester].

Objective: To analyze the clinical features and to explore the etiology of short fetal femur during the third trimester. Methods: From January 2010 to June 2016, 21 singleton pregnancies with short fetal femur detected by ultrasonography during the third trimester were referred to the Chinese PLA General Hospital. Clinical data were collected, karyotype or single nucleotide polymorphism microarray was carried out to detect chromosomal abnormalities, and FGFR3 c.1138G>A mutation detection was carried out to detect achondroplasia (ACH) via invasive procedure, respectively. The deviation of femur length from the mean value of the gestational age in ultrasonography was expressed as the Z-score. The difference between ACH and isolated short femur (ISF, in the absence of associated structure abnormality or genetic abnormality) was then explored. Results: In the 21 fetuses, 11 had abnormal genetic test results(52%, 11/21), including 9 cases of ACH, 1 case of Ellis-van Creveld Syndrome and 1 case of Pallister-Killian syndrome. In the 10 ISF fetuses (48%, 10/21), 3 cases were fetal growth restriction, 1 was normal small for gestational age infant and 6 cases were unexplained. The median Z-scores for 9 cases of ACH and 10 cases of ISF in the third trimester were -5.04, -3.20, respectively. The short femur in ACH was more severe than in ISF (P=0.005) in the third trimester. Conclusions: The etiology of short fetal femur is complicated, including skeletal dysplasia, chromosomal abnormality, fetal growth restriction, as well as normal variants during fetal development. Genetic test should be considered during the antenatal consultation.

Association of single-nucleotide polymorphisms, rs2235371 and rs2013162, in the IRF6 gene with non-syndromic cleft palate in northeast China.

The aim of this study was to determine the association between two SNPs (rs2235371 and rs2013162) in the interferon regulatory factor 6 (IRF6) gene and non-syndromic cleft palate (NSCP) in northeast China. We genotyped these two SNPs in 104 NSCP cases, as well as in 178 parents and 300 controls. Case-control and case-parent analyses were performed using χ2 tests and family-based association tests (FBAT). Results indicated that there were significant differences in both genotypic and allelic distributions between patients and controls at rs2235371 and rs2013162 in the IRF6 gene. Case-parent analysis revealed over-transmission of the C allele in rs2235371 and the A allele in rs2013162. Lastly, FBAT showed over-transmission of the CA haplotype. This study demonstrated that the two SNPs, rs2235371 and rs2013162, are strongly associated with NSCP in the northeast Chinese population.

[The efficacy of external physical vibration lithecbole in the assistant treatment of the upper urinary tract residual stones: a meta-analysis].

Objective: To determine the clinical effect of Friend-Ⅰ External Physical Vibration Lithecbole (EPVL) by Meta-analysis. Methods: Pubmed, Embase, Medline, Cochrane Library, Chinese National Knowledge Infrastructure were searched for clinical trials comparing EPVL with the conventional treatment. The quality of included studies was assessed and Meta-analysis was conducted by Review Manager 5.3 software. Results: Five randomized or Quasi-randomized controlled trials met the inclusion criteria. The first day stone expulsion rate of EPVL group was superior to the control group (OR=4.95, 95% CI: 3.35-7.32, P<0.000 01). Both one-week (OR=3.13, 95% CI: 1.95-5.04, P<0.000 01) and two-week stone free rate (OR=4.50, 95% CI: 2.02-10.00, P=0.000 2) were statistically higher in the EPVL group than that in the control group. No severe adverse event occurred during the follow-up. Conclusions: Our study suggested that EPVL could be the effective treatment for upper urinary tract residual stone. However, more high quality randomized controlled trials are needed to better affirm this.

Variations in WNT3 gene are associated with incidence of non-syndromic cleft lip with or without cleft palate in a northeast Chinese population.

Non-syndromic cleft lip with or without cleft palate (NSCL/P) is a common birth defect. Several WNT genes are involved in craniofacial embryogenesis, and therefore may play an important role in the etiology of NSCL/P. Two SNPs (rs3809857 and rs9890413) in the WNT3 gene were subjected to case-control and case-parent analysis by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) in 236 unrelated patients with NSCL/P, including 128 elementary families (185 mothers and 154 fathers), and 400 control individuals from northeast China. The rs3809857 SNP, under the assumption of a dominant model, was found to induce a 2-fold lower risk of NSCL/P ORGG vs GT + TT = 0.605, 95%CI = 0.436-0.839, P = 0.003). Moreover, the family-based association test revealed an under-transmission for the minor allele T. On the other hand, we observed a significant association in the case-control and case-parent analysis of the SNP rs9890413. In addition, the P values for the haplotype of rs3809857-rs9890413 were observed to be statistically significant (P = 0.004). In conclusion, our study confirmed the association between the WNT3 variant and NSCL/P in the population tested.

Mutation analysis of four Chinese families with pure hereditary spastic paraplegia: pseudo- X-linked dominant inheritance and male lethality due to a novel ATL1 mutation.

We studied four Chinese families with pure hereditary spastic paraplegia (HSP) to investigate the clinical features and associated genetic mutations. Linkage analysis was performed for all families to map the disease locus onto autosomal chromosomes, and related loci involved in HSP on the X chromosome were also examined. Polymerase chain reaction (PCR) sequencing was used to detect gene mutations. To confirm the influence of a splice-site mutation on mRNA, we used reverse transcription-PCR and direct sequencing. Linkage analysis and ATL1 gene sequencing of amniocytes were performed for prenatal genetic diagnosis. One missense variant (c.1517T>A) and a splice-site mutation (c.1245+1G>A) in SPAST, and two missense variants (c.715C>T, c.1204T>G) in ATL1 were identified. The c.1245+1G>A mutation caused a deletion of exon 9 in the SPAST gene. Prenatal genetic diagnosis showed that fetus did not carry the ALT1 c.1204T>G mutation. Follow-up was maintained for 5 years, and the negative result was confirmed by evidence of a healthy growing boy. We identified two novel mutations and two previously reported mutations in SPAST and ATL1, respectively. The family with the ATL1 c.1204T>G mutation exhibited male-lethality, female infancy-onset, and pseudo- X-linked dominant transmission, which had never been previously reported for HSP. Characteristic facial features were also noticed. The boy on whom prenatal gene diagnosis was performed is healthy and without unusual facies, suggesting that the c.1204T>G mutation might be related to these features. The results extend the genetic spectrum of HSP and suggest that linkage analysis remains a powerful tool in gene discovery studies.

Study on the electro-transformation conditions of improving transformation efficiency for Bacillus subtilis.

AIMS: To optimize the transformation conditions and improve the transformation efficiency of Bacillus subtilis WB800 and DB104. METHODS AND RESULTS: Trehalose, which could decrease the damage of electric shock to the cells, was added to the electroporation medium containing sorbitol and mannitol. The factors affecting the transformation efficiency, such as the growth phase of bacteria, cell concentration, electric field strength and plasmid variety, were examined and improved. The new method increased the transformation efficiency of B. subtilis by nearly 100-fold compared with the conventional one. CONCLUSIONS: With the optimized method, the transformation efficiency came up to 3.64 × 10(5) transformants µg(-1) DNA for WB800, and 2.10 × 10(5) transformants µg(-1) DNA for DB104. SIGNIFICANCE AND IMPACT OF THE STUDY: This improvement in transformation efficiency will be largely attributed to the research of expression of exogenous genes in B. subtilis, gene library construction for directed evolution and transformation of wild-type B. subtilis strains.

Fine mapping of a gene for non-pollen type thermosensitive genic male sterility in rice (Oryza sativa L.).

The thermo-sensitive genic male sterility (TGMS) lines play a crucial role in two-line hybrid rice production. For a practical TGMS line, the stability of male sterility is one of the most important technical indicators. In this study, XianS, a spontaneous mutant with stable male sterility from an indica rice cultivar Xianhuangzhan, was classified as a non-pollen type TGMS line. The critical non-pollen sterility point temperature of XianS was determined as 27 degrees C. Genetic analysis demonstrated that the non-pollen sterility in XianS was controlled by a single recessive gene. Using SSR markers and bulked segregant analysis, the TGMS gene in XianS was fine mapped to a 183 kb interval between RMAN81 and RMX21 on chromosome 2. Two markers, 4039-1 and RMX14 completely cosegregated with this gene. Allelism test indicated that the non-pollen phenotype in seven non-pollen type TGMS lines from different sources, XianS, AnnongS-1, Q523S, Q524S, N28S, G421S, and Q527S is caused by the same TGMS gene. Although the location of TGMS gene in XianS is close to the gene OsNAC6, a previously identified candidate gene of tms5 in AnnongS-1, the sequence of OsNAC6 and its promoter region was identical in TGMS line XianS, AnnongS-1, and wild-type Xianhuangzhan. These results suggest that the non-pollen type TGMS trait probably be controlled by the same TGMS gene in different TGMS rice lines, but its real candidate gene still need to be further studied and identified.

Terahertz response in single-walled carbon nanotube transistor: a real-time quantum dynamics simulation.

We use time-dependent quantum wavepacket methods to simulate ballistic electron transport in a single-walled carbon nanotube field-effect transistor at terahertz frequencies ( approximately 100 GHz-10 THz). We observe an electron resonance phenomenon in a sub-picosecond-scale time domain. Our simulation results clearly show that the electron resonance corresponds to the formation of the resonance cavity and the interference of the electron wavepackets, which is directly supported by recent experimental measurements (Zhong et al 2008 Nat. Nanotechnol. 3 201).

Inhibition of connective tissue growth factor by small interfering RNA prevents renal fibrosis in rats undergoing chronic allograft nephropathy.

AIM: Connective tissue growth factor (CTGF) is a highly profibrogenic molecule implicated in renal fibrogenesis. Small interfering RNA (siRNA) is an effective tool to silence gene expression. This study determined whether caudal vein injection of siRNA targeting CTGF inhibited its expression in rat kidneys in vivo, and furthermore whether it protected the kidney from renal fibrosis in chronic allograft nephropathy (CAN). METHODS: Male inbred Fischer (F344, RT1(lv1)) rat renal grafts were orthotopically transplanted into Lewis (LEW, RT1(1)) rats following the procedure of Kamada with our modification. At 6 weeks, recipients were divided into siRNA, normal saline (NS), and control siRNA groups, using daily siRNA-targeting CTGF (0.1 mg/kg), or NS, or a control siRNA via caudal vein injection for 14 days. At 4, 6, and 8 weeks, we observed the pathologic changes, expression of CTGF, E-cadherin, collagen I and IV, and anti-smooth muscle actin (alpha-SMA). RESULTS: Serum creatinine level, Banff score, and the expression of CTGF were significantly lower among the siRNA than the NS or the control siRNA groups at 8 weeks (P < .05). The expressions of collagen I and IV, and alpha-SMA were also significantly downregulated and E-cadherin was lost in the siRNA versus the NS and control siRNA groups at 8 weeks. CONCLUSIONS: This study showed that delivery of CTGF siRNA via the caudal vein significantly inhibited expression of CTGF in rat kidneys, effectively preventing fibrosis in CAN. The results suggest that siRNA-targeting of CTGF has the potential to be a novel strategy for amelioration of CAN.

C4d deposition is associated with chronic allograft nephropathy in rats and could be influenced by immunosuppressants.

AIMS: Deposition of C4d in peritubular capillaries (PTC) has been considered to be a marker of humoral immunity in renal transplant. This study is to investigate C4d deposition in rat renal allografts undergoing CAN and the effects of immunosuppressants on it. METHODS: Fisher 344 rat renal grafts were orthotopically transplanted into Lewis rats following the procedure of Kamada with our modification. All the recipients were given CsA 10 mg/kg(-1).d(-1) x 10 d and then divided into 5 groups (each n = 9); (1) Vehicle: vehicle orally, (2) CsA: 6 mg/kg(-1).d(-1), (3) RAPA: 0.8 mg/kg(-1).d(-1), (4) FK 506: 0.15 mg/kg(-1).d(-1), (5) MMF: 20 mg/ kg(-1).d(-1). At 4 weeks, 8 weeks, 12 weeks, the rats were sacrificed, renal allografts were harvested and sera were collected. The deposition of C4d was detected by immunofluorescence and analyzed by Integrated Optical Density (IOD). The pathological changes were accessed according to the Banff 97 criteria. RESULTS: C4d deposition in PTC was found in all the allografts at 4 weeks, while there was no obvious manifestations of CAN in all the groups; the differences of Banff Score between all groups were not significant (P > .05). The values of IOD in RAPA and MMF group were lower than those in other 3 groups (P = .002, .006). The differences between RAPA and MMF, and between other 3 groups were not significant (P > .05). The intensity of C4d increased along with the progression of CAN, the heaviest C4d deposits in PTC were found at 12 weeks, and meanwhile the severest CAN was found. Comparing with Vehicle group, CsA and FK 506 had no effect on C4d deposition (P > .05), however, MMF and RAPA obviously decreased the C4d deposition (P = .000). The intensity of C4d deposition had a significant correlation with the severity of CAN (r = 0.894, P = .000). CONCLUSIONS: Our study suggests that the deposition of C4d in allografts appears earlier than pathological changes of CAN and has a correlation with the progression of CAN. MMF and RAPA can attenuate CAN by inhibiting humoral immunity. In contrast, CsA and FK 506 have no effect on humoral immunity.

C4d deposition is correlated with the level of antivimentin antibody in rat kidneys undergoing chronic allograft nephropathy.

AIMS: Antivimentin antibody is often produced as an autoantibody after transplantation. C4d deposition, a marker of humoral immunity during transplantation, is believed to reflect alloantibodies. This study investigated the relationship between C4d deposition and humoral immunity to vimentin among rat kidneys undergoing chronic allograft nephropathy (CAN). METHODS: Fisher 344 rat renal grafts were orthotopically transplanted into Lewis rats following the procedure of Kamada with our modification. All recipients were administered cyclosporine (CsA) (10 mg/kg(-1).d(-1) x 10 d) before being divided into 3 groups of oral treatments: (1) vehicle, (2) CsA (6 mg/kg(-1).d(-1)), and (3) mycophenolate mofetil (MMF; 20 mg/kg(-1).d(-1)). At 4, 8 and 12 weeks after transplantation, the rats were killed, the renal allografts harvested, and the sera collected. Serum creatinine (SCr) was measured and pathologic changes assessed according to the Banff 97 criteria. The antivimentin antibody was quantified by enzyme-linked immunosorbent assay. The deposition of C4d detected by immunofluorescence was analyzed by integrated optical density (IOD). RESULTS: Antivimentin antibody was observed in sera of all transplanted rats. The level of antivimentin antibody (IgGDeltaOD) increased gradually during the development of CAN from 4 weeks. Simultaneously, C4d deposition in peritubular capillaries also progressively strengthened. There was a strong positive correlation between the content of antivimentin antibody and C4d deposition (r = 0.892; P = .000). MMF simultaneously decreased antivimentin antibody formation and C4d deposition. In contrast, CsA had no significant effect. CONCLUSIONS: We demonstrated the production of antivimentin antibodies and the deposition of C4d during the development of CAN. There was a positive correlation between them. Whether humoral immunity to vimentin contributes to C4d deposition is not clear and further studies are needed to elucidate this issue.

Expressions of Angiopoietin-1, Angiopoietin-2, and Tie2 and their roles in rat renal allografts with chronic allograft nephropathy.

OBJECTIVE: Angiopoietin-1 (Ang1) and -2(Ang2) are 2 ligands for the endothelium-specific tyrosine kinase Tie2. Previous studies have shown that reciprocal regulation of Ang1, Ang2, and Tie2 plays an important role in chronic cardiac allograft vasculopathy. This study investigated the expressions of Ang1, Ang2, and Tie2 in rat renal allografts undergoing chronic allograft nephropathy (CAN). MATERIALS AND METHODS: Renal transplantations following the procedure of Kamada with our modification were orthotopically performed using Fisher (F344, RT1(1v1)) rats as both donors and recipients in the autograft group. Fisher and Lewis (LEW, RT1(1)) rats were used as donors and recipients in the allograft group, respectively, which was treated with cyclosporine (CsA; 10 mg/kg/d x 10 d). At 4w, 8w, and 12 weeks posttransplantation, serum creatinine (SCr) was measured and pathologic changes assessed according to the Banff 97 criteria. The mRNA (Deltact) and protein expressions of Ang1, Ang2, and Tie2 were localized by real-time fluorescence quantitative polymerase chain reaction (PCR) and by immunohistochemistry. RESULTS: The elevation in SCr and the pathologic changes in CAN were observed in all allografts at 8 and 12 weeks. The expressions of Ang1 and Ang2 were localized to epithelial cells and endothelium of the vascular bundles of the glomeruli; Tie2 was specifically expressed in endothelium of vessels both in auto- and allografts at all time points posttransplantation. At 4 weeks, the differences in mRNA expression of Ang1, Ang2, and Tie2 between the 2 groups were not significant (P > .05). Compared with autografts, the mRNA expression of Ang1 decreased significantly (P = .008 and .003 for 8 and 12 weeks, respectively), and the mRNA expressions of Ang2 and Tie2 significantly increased (P = .001/.006 and .005/.001 for 8 and 12 weeks, respectively). The changes in expression of all 3 genes showed significant correlation with the Banff score in the allografts. CONCLUSION: This study suggested that the abnormal expression and reciprocal regulation of Ang1, Ang2, and Tie2 may play important roles in the development of CAN in rat renal allografts.

Epithelial to mesenchymal transformation in tubular epithelial cells undergoing anoxia.

AIM: Epithelial-mesenchymal transformation (EMT) has been proved to be a critical event in fibrogenesis of renal allografts. This study sought to determine whether anoxia could induce EMT from tubular epithelial cells (TEC). METHODS: Rat TEC-line (NRK-52E) was cultured in Dulbelco's modified Eagle's medium (DMEM) without glucose under 100% N2 for 4 hours. After 6, 12, 24, 48, and 72 hours, the expressions of connective tissue growth factor (CTGF) mRNA and protein were measured by real-time RT-PCR and Western blot, respectively. Morphologic changes and cytoskeleton remodeling were observed in NRK-52E cells under laser confocal microscopy. Immunohistochemistry and flow cytometry were used to detect expression changes of E-cadherin, alpha-smooth muscle actin (SMA), types I and IV collagen, all of which are involved in TEC, EMT. RESULTS: After stimulation by anoxia, NRK-52E cells became round and enlarged with a remodeled cytoskeleton. The expressions of CTGF mRNA and protein were upregulated after 6 hours, reaching their peak at 48 hours. The expressions of types I and IV collagen, and alpha-SMA were all upregulated except for E-cadherin. CONCLUSIONS: Anoxia upregulated the expression of CTGF and other EMT-associated genes in NRK-52E cells.

Rapamycin and cyclosporine have different effects on expression of Ang-1 and Ang-2 and Tie2 in rat renal allograft with chronic allograft nephropathy.

OBJECTIVE: Previous studies have indicated that angiopoietin-1 (Ang1) and angiopoietin-2 (Ang2) and tyrosine kinase receptor Tie2 regulate the maintenance and integrity of blood vessels and have potential anti-inflammatory properties. This study of cardiac allografts investigated whether there is a difference between rapamycin and cyclosporine A (CsA) in the ability to affect expression of Ang1, Ang2, and Tie2 in rat renal allografts with chronic allograft nephropathy (CAN). METHODS: A male inbred F344 to Lewis rat renal CAN model was established via a modified Kamada procedure. The recipients were first treated with CsA, 10 mg/kg/d, for 10 days and then allocated randomly to three oral treatment groups: control; CsA, 6 mg/kg/d; and rapamycin, 0.8 mg/kg/d. At 4, 8, and 12 weeks posttransplantation, the rats were killed to harvest the renal allografts. The serum creatinine concentration (SCr) was measured, and the pathologic changes were assessed according to Banff 97 criteria. The expression of messengerRNA and proteins of Ang1, Ang2, and Tie2 was determined using real-time fluorescence quantitative polymerase chain reaction and immunohistochemistry. RESULTS: The elevation of SCr and the pathologic changes of CAN were observed in the control and CsA groups at 8 and 12 weeks; the differences between the 2 groups were not significant (P > .05). The levels of SCr and Banff score in the rampamycin group were lower than those in other 2 groups (P < .01). The expression of Ang1 and Ang2 was localized to epithelial cells and endothelium of vascular bundles of glomeruli, and Tie2 was specifically expressed in the endothelium of vessels in all 3 groups. At 4 weeks, the differences in mRNA expression of Ang1, Ang2, and Tie2 between 3 groups were not significant (P > .05). In a comparison of the control and CsA groups, mRNA expression of Ang1 was increased (P < .05), and mRNA expression of Ang2 and Tie2 was decreased (P < .05) in the rapamycin group at 8 and 12 weeks. The differences between the control and CsA groups were not significant at 8 or 12 weeks (P > .05). CONCLUSIONS: Our results show that compared with CsA, rapamycin modulates the expression of Ang1, Ang2, and Tie2 in rat renal allografts with CAN, which suggests that rapamycin may improve the long-term survival of renal allografts through its vasculoprotective properties.

Effects of mycophenolate mofetil on chronic allograft nephropathy by affecting RHO/ROCK signal pathways.

AIM: We sought to investigate the effects of mycophenolate mofetil (MMF) on chronic allograft nephropathy (CAN) by affecting Rho and ROCK signal pathways. METHODS: Male inbred F344 rat renal grafts orthotopically transplanted into Lewis rats were first treated with CsA (10 mg/kg(-1).d(-1) x 10 d) and then divided into 3 groups (each n = 9): (1) orally vehicle, (2) cyclosporine (CsA, 6 mg/kg(-1).d(-1)) and (3) MMF (20 mg/kg(-1).d(-1)). In addition we performed autografts of F344 (n = 10) at 4, 8, and 12 weeks, serum creatinine (SCr) was measured and pathologic changes assessed. Expression of RhoA and ROCK-1 was determined by real-time reverse transcriptase polymerase chain reaction. Expressions of alpha-smooth muscle actin (alpha-SMA) and connective tissue growth factor (CTGF) were observed by immunohistochemistry. RESULTS: SCr and Banff score began to increase at 4 weeks in all 3 allografted groups with obvious deterioration in both the vehicle and CsA groups at 8 and 12 weeks. The differences between vehicle/CsA and autografts were significant (P = .000). SCr and Banff score among the MMF group increased mildly and moderately at 8 and 12 weeks, respectively, but were significantly lower than those in the vehicle/CsA cohort (P < .05). Expressions of RhoA and ROCK-1 mRNAs and proteins were observed in mesangial and tubular cells, increasing gradually along with the progression of chronic allograft nephropathy (CAN). There was a negative correlation between RhoA/ROCK-1 mRNA and Banff score (r = -.637, p = .000; r = -.676, P = .000) or SCr (r = -.705, P = .000; r = -.756, P = .000). MMF downregulated gene and protein expressions of RhoA and ROCK-1. CsA had little effect on these expressions. Expressions of alpha-SMA and CTGF were observed in renal epithelial and tubular cells. CONCLUSION: Herein we have demonstrated abnormal expression of RhoA and ROCK-1 signal pathways, which may play roles in CAN. MMF may attenuate CAN by downregulating the expression of RhoA/ROCK-1, alpha-SMA, and CTGF.

Antidepressant effects of Kai-Xin-San in fluoxetine-resistant depression rats.

This study aimed to investigate the antidepressant effect and the mechanism of action of Kai-Xin-San (KXS) in fluoxetine-resistant depressive (FRD) rats. Two hundred male Wistar rats weighing 200±10 g were exposed to chronic and unpredictable mild stresses (CUMS) for 4 weeks and given fluoxetine treatment simultaneously. The rats that did not show significant improvement in behavioral indexes were chosen as the FRD model rats. These rats were randomly divided into four groups: FRD model control; oral fluoxetine and aspirin; oral KXS at a dose of 338 mg·kg-1·day-1; and oral KXS at a dose of 676 mg·kg-1·day-1. Rats continued to be exposed to CUMS and underwent treatment once a day for 3 weeks, then cytokine (COX-2, IFN-γ, IL-1ß, IL-2, IL-4, IL-6, IL-10, TGF-ß, and TNF-α) levels in the hippocampus and serum, and organ coefficients were measured. Both doses of KXS improved the crossing and rearing frequencies, sucrose-preference index, and body weight in FRD rats. KXS at a dose of 338 mg·kg-1·day-1reduced COX-2, IL-2, IL-6, TNF-α levels, increased IL-10 level in the hippocampus, and reduced IL-2 and TNF-α levels in serum. KXS at a dose of 676 mg·kg-1·day-1reduced TNF-α level in the hippocampus, reduced IL-2 and TNF-α levels in serum, and increased IFN-γ and IL-10 levels in the hippocampus and serum. There were no significant differences in organ-coefficients of the spleen among and between groups. The results suggested that oral administration of KXS in FRD rats was effective in improving behavior disorders by influencing various inflammatory pathways.