OBJECTIVETo observe the effects of surgical delay procedure on the survival of perforator flap with three angiosomes in rat, and to explore its possible mechanism.

METHODSThe flap model was a perforator flap with three angiosomes which located on the right dorsal side of a rat based on the right deep circumflex iliac vessel. The two connection areas between the three angiosomes were successively named choke zone (CZ) 1 and CZ 2 beginning from the pedicle to the remote area. A total of 110 SD rats were divided into routine flap group (RF, n = 40), delay only group (DO, n = 30), and delay flap group (DF, n =40) according to the random number table. (1) In group RF, 30 rats were selected according to the random number table, and flap surgery was performed directly. Six rats were sacrificed on post operation day (POD) 0, 1, 2, 3, 7 respectively to collect the full-thickness skin samples at both CZs for HE staining to measure the vascular density and diameter. The rest 10 rats underwent flap surgery immediately after a catheter was successfully implanted into their external jugular vein. A volume of 1.5 mL sodium fluorescein solution (100 g/L) was injected to the 10 rats on POD 0 (5 rats) or POD 1 (5 rats) each time with a 2-day interval to learn the change in flap circulation. Each rat was injected for 4 times. The flap survival rate of the 10 rats was calculated on POD 7, and the configuration and distribution of the vessels in the flap were observed through angiography with the improved perfusion method of lead oxide-gelatin. (2) In group DO, the right thoracodorsal perforators of all the rats were surgically ligated through a small skin incision, and 6 rats were sacrificed on POD 0, 1, 2, 3, 7 respectively. The skin samples of each rat at the same area as in group RF were harvested to measure the vascular density and diameter. (3) In group DF, rats were treated with ligation surgery as in group DO, and then they were assigned and treated as in group RF on POD 7 with corresponding indexes detected later. Data were processed with group t test, analysis of variance with factorial design, and SNK test.

RESULTS(1) Significant differences of vascular density at both CZ 1 and CZ 2 were found on POD 7 among the three groups ( with F values respectively 2. 69 and 2. 76, P values below 0.05). The vascular density values of CZ 1 and CZ 2 of rats in group DF were (29 ± 7) and (31 ± 8) per mm on POD 7, which were significantly higher than those of group RF [(23 ± 5) and (23 ± 3) per mm2, with q values respectively 5.67 and 6.01, P values below 0.05] and those within group DF on POD 0 (with q values respectively 6.42 and 7. 14, P values below 0. 05). On POD 3 and 7, the vascular diameter values of CZ 1 of rats in groups RF and DF were significantly higher than those of group DO (with q values from 8. 15 to 11.13, P values below 0.05). The vascular diameter values of CZ 2 of rats in group DF onPOD 0, 1, 2, 3,7 [(65 ± 8), (63 ± 13), (69 ± 9), (67 ± 8), (64 ± 13) 230m] and in group DO on POD 3 and 7 were significantly higher than those in group RF [respectively (46 ± 10) , (40 ± 9), (43 ± 13), (46 ± 12), (47 ± 11) µm on POD 0, 1,2, 3, 7 ] at corresponding time point (withqval- ues from 7.29 to 10.79, P values below 0.05). The difference in vascular diameter between CZ 1 and CZ 2 was statistically significant in groups RF and DO on POD 3 and 7, and in group DF on POD 0, 1 , and 2 (with q values from 5.32 to 9.56, P values below 0.05). Compared with that on POD 0 within each group, the vascular diameter of CZ 1 in groups RF and DF and that of CZ 2 in group DO increased significantly on POD 3 or 7 (with q values from 6.12 to 8.13, P values below 0.05). (2) In groups DF and RF, blood from the pedicle ran through CZ 1 and covered the dynamic territory successfully within POD 7. On POD 0, the blood within all flaps was blocked for about 3 min after going through CZ 1 at 1 cm distal from CZ 2 in group DF and around CZ 2 in group RF. (3) Flap survival rate of rats in group DF was (95 ± 12) % , which was statistically higher than that of group RF [(80 241 9) % , t = 2.91, P <0.01]. All the partial flap necrosis occurred in potential territory. (4) Compared with the vessels in the left dorsal side without surgery, the vessels of CZ 1 in group RF were dilated obviously, and the boundary between vascular trees became indistinct, but the vessels in CZ 2 changed slightly; the vessels in both CZs in group DF were dilated dramatically.

CONCLUSIONSThe delay method could enhance the survival of potential territory in perforator flap with three angiosomes, and it acted mainly by dilating the choke vessels in CZ 2 before flap surgery.

Angiography ; Animals ; Graft Survival ; physiology ; Male ; Necrosis ; Perforator Flap ; blood supply ; physiology ; Rats ; Skin ; blood supply ; Surgical Flaps ; blood supply ; physiology ; Time Factors

OBJECTIVETo develop a new experimental animal model of different a single perforating vessel as its pedicle, and to investigate this vessel can captures how many adjacent angiosomes in different directions.

METHODSThirty-six Sprague-Dawly rats of both sexes were used. The rats were divided into group A, group B and group C. Group A: the unilateral deep circumflex iliac perforator artery- based flap. Group B: the unilateral posterior intercostal perforator artery-based flap. Group C: the unilateral lateral thoracic perforator artery-based flap. An extended dorsal perforator flap measuring up to 13 cm x 6 cm was designed in 36 rats to assess the viability of the flap. The upper margin was located at the level of the tip of the scapula and the lower margin at a level 1 cm below the iliac crest. All flaps were observed for 7 days postoperatively, 72 hours after flap elevation, observe flap dyeing conditions through the vivo fluorescein injection, the surviving flap area was calculated as a percentage of total flap dimensions and the angiosome's structure of the flap was displayed by radiopaque microangiography.

RESULTSNo fluorescence was visible in the distal flap of groups A and C, the whole flap show bright fluorescence in group B. Survival rate of C, A, B were improved in order. Statistic difference is significant (P < 0.01) between group and group. In group A, lead oxide-gelatin angiography shows the cephalic flap necrosis occurred in the bilateral lateral thoracic territories, and the vascular architecture partly disappeared in the necrotic area. In group B, the vascular architecture of flap is unbroken. In group C, the caudal flap necrosis occurred in the bilateral deep circumflex iliac perforator artery territories, and the vascular architecture partly disappeared and disordered in the necrotic area.

CONCLUSIONSThe perforator flap is based centrally on a single perforator, this vessel can capture multiple the second vascular territory. In a direction, the longest distance that the blood supply can reach is the point of the third perforator vessel puncture into skin, which can provide certain theoretical guidance for designing of perforator flap.

Angiography ; Animals ; Female ; Graft Survival ; Male ; Models, Animal ; Perforator Flap ; blood supply ; Rats ; Rats, Sprague-Dawley

AIM: To explore the molecular mechanism of asthenospermia(AST) by preliminary screening of nucleotide sequences from the ND3 and ND4L genes of mitochondrial DNA(mtDNA). METHODS: Samples from 50 AST patients and 42 age-matched normal controls were collected according to the WHO criteria. Density gradient centrifugation was applied to separate spermatozoa with different vigor. The ND3 and ND4L genes of mtDNA were amplified and sequenced directly from the extracted genomic DNA from AST patients and normal controls. The sequences were compared with revised Cambridge Reference Sequence(rCRS) to analyze the variants. RESULTS: A total of 22 nucleotide variations were found in ND3 and ND4L genes of mtDNA in asthenospermia group and control group. G10320A, A10398G and T10609C were missense mutations, while A10157G and A10313C were the reported for the first time in this study. Haplotype N in patients with AST(33/50) was higher than that in control group(14/42, P<0.05), and haplotype R9 in patients with AST(15/50) was also higher than that in control group(4/42, P<0.05) through genetic testing of ND3 gene. Rates of sperm progressive motility of haplotype F1, F2 and R9 were significantly lower than those of haplotype M and M rest. Two haplotype differences, haplotype M and N, were found in the same AST patient's spermatozoas which had different vigor. Haplotype M had stronger vigor, while haplotype N had lower vigor. By sequencing ND3 gene of mtDNA from 50 AST patients, we detected G10310A heteroplasmic mutation in 2 specimens of asthenospermia with poor and moderate motility spermatozoa, respectively. No mutation occurred in good motility spermatozoa. CONCLUSION: Haplotype of mitochondrial may have some correlation with sperm motility. The nt10398G-10400T polymorphisms may have benefit for sperm motility, whereas the mutation in nt10310A may impair sperm motility.

Objective To investigate the relationship between mtND4 point mutation in sperms and asthenospermia. Methods Fifty-six asthenospermia cases and 44 control cases were collected using the WHO criterion for defining asthenospermia, the regions of mtND4 gene were amplified by using PCR of 3 pairs primers. Consequently, the point mutation, missense mutation and multiple single nucleotide polymorphisms (SNP) were analyzed by employing sequencing technology and bioinformatics tools. Results Six mutations never before identified were found. The frequency of single point mutation T10873C and T11944C in the control group were significantly higher than those in the asthenospermia group (P＜0.05). Eight cases involved T10873C or T11944C among the 10 cases in control groups with missense mutations were found. But, there were only 2 cases with such mutation in the 10 asthenospermia cases with missense mutations (P＜0.05). The previous 20 cases of missense mutations can be described as either multiple SNP group (with T10873C or T11944C) or nonmultiple SNP group. The percentage of a range and a plus b range of multiple SNP group of sperm was significantly higher than the non-multiple SNP group(P＜0.05). Conclusions mtND4 gene mutation, especially the missense mutation may induce loss of sperm motility. The mutations of T10873C and T11944C may be useful for sperm motility or counteract the influence for the sperm motility caused by these harmful mutations.

Objective To construct the recombinant plasmid containing human filaggrin gene,purify and identify the immunoreactivity of the recombinant protein,and establish the indirect ELISA to detect AFA for diagnosis of RA.Methods The constructed plasmids were transformed into E. Coli Rosettagami(DE3).This fusion protein was purified by NAT chromatography.ELISA coated with the fusion protein Was established to detect the AFA in serum of patients,which included 114 cases of RA,56 cases of SLE,32 cases of OA and 40 cases of normal controls. The correlation between the results of AFA and anti-CCP in RA group were compared. Results 321 bp fragment of filaggrin gene was amplified and the recombinant expression vector pET-28a( + )-filaggrin was constructed. The sequence of filaggrin gene was the same as the sequence reported in the literatures. The Rosetta-gami (DE3) strains of E. Coli with recombinant vector showed high level of filaggrin protein after induction. The SDS-PAGE showed that the plasmid expressed the filaggrin fusion protein with molecule weight of 14 000 Da. The expression protein could be purified by Ni-NAT with activity. The absorbance value of AFA in RA group was 0.473 ±0. 248 while they were 0. 160 0. 088, 0. 121±0. 070, 0.050 0. ±018 in SLE, OA and normal groups respectively. There were significant differences of absorbance values of AFA between RA and SLE, OA, control group (t = 12.004, 14. 464, 18.078, P<0. 01, respectively). The positivities of anti-filaggrin in RA, SLE and OA were 48.2%, 5.4% and 3. 1% respectively. The positivities of AFA were significantly different between RA, OA and normal control groups (x~2 = 67. 088, P < 0. 01). There was positive correlation of results between AFA and anti-CCP antibody (r = 0.42, P < 0. 05 ) . The consistency rate of results between AFA and anti-CCP was 70. 1%. Anti-CCP was negative in 10 out of 114 patients with AFA positive. AFA can be used to diagnose RA with sensitivity of 48. 2% , specificity of 96.9% , positive predictive value of 93. 2% and negative predictive values of 67. 9% . Conclusions The purified human filaggrin fusion protein is successfully purified. The indirect ELISA method based on the recombinant protein shows good sensitivity and specificity. Joint detection with AFA and anti-CCP can improve the positive rate of detection.

BACKGROUND:Knee osteoarthritis is a chronic disease characterized as cartilage degeneration and hyperostosis to impact the soft tissues around the joints, thereby resulting in joint pain, and it can be aggravated and lead to joint deformity after weight-bearing. OBJECTIVE:On the basis of imaging parameters of knee osteoarthritis patients and normal controls, a model was established. And then, knee squatting movement was simulated on the model and stress data from the cartilage surface of patients and normal controls were colected and compared, thereby to obtain the stress characteristics of the cartilage surface in knee osteoarthritis patients. METHODS: There were 30 knee osteoarthritis patients and 30 healthy volunteers in the study. CT and MRI data from these participants were colected and analyzed using Mimics software and simpleware software to establish a model. Based on this model, cartilage-related stress data were harvested from knee osteoarthritis patients and healthy volunteers and compared to summarize the stress characteristics of the articular cartilage in patients with knee osteoarthritis. RESULTS AND CONCLUSION:In knee osteoarthritis patients, the stress on the cartilage surface from standing to squatting was increased in a nonlinear manner. During the process of squatting, the mediolateral stress was higher in the knee osteoarthritis patients than healthy controls (P < 0.05); however, there was no difference in the proximodistal and anteroposterior cartilage surface stress between the two groups (P > 0.05). These findings indicate that the knee osteoarthritis musculoskeletal model and finite element model are established successfuly, and these models are closer to the real motion characteristics of the knee joints, based on which, quantitative biomechanical data of the knee can be provided for knee osteoarthritis treatment.

OBJECTIVEBy bioinformatics method, the effect in hematopoietic system of bioactive peptide HP-6, which was obtained from donkey serum albumin and is one of the major protein components from donkey-hide gelatin, was investigated.

METHODHuman bone marrow nucleated cells (hBMNCs) and murine bone marrow stromal cells (mBMSCs) were separated and cultured with different concentration of peptide HP-6 (0.000 15, 0.001 5, 0.015, 0.15, 1.5 micromol x L(-1)). The effect on promoting proliferation of cells related to hematopoiesis in bone morrow was detected and the ultrastructure of cells after treated by HP-6 was observed through transmission electron microscope. Hemorrhage anemia mouse model and anemia mouse model induced by cyclophosphamide were established, and randomly divided into peptide HP-6 groups which were administered respectively with different doses (1, 0.1, 0.01 mg x kg(-1)) by gavage, and control group which was administered with PBS by gavage. Peripheral blood components of all mice and bone morrow cells (BMC) number of mice induced by cyclophosphamide were evaluated.

RESULTPeptide HP-6 could concentration-related promote the proliferation of hBMNCs and mBMSCs, hBMNCs got the highest reproduction rate of 152.11% and mBMSCs also got 63.52% with the concentration of 0.15 micromol x L(-1), then the reproduction rate decreased while the concentration kept increasing. The transmission electron microscope showed that ultrastructure of cells was normal after treated by HP-6.1 mg x kg(-1) peptide HP-6 significantly increased peripheral platelet and protected mouse morrow injured by cyclophoshamide. 0.1 mg x kg(-1) peptide HP-6 significantly increased peripheral platelet with relative growth rate of 77.65%, increased peripheral white blood cells count and peripheral red blood cells count, also could protect mouse peripheral blood after treated by chemotherapeutics.

CONCLUSIONPeptide HP-6 could promote the proliferation of cells related to hematopoietic system, enhance mouse hemopoiesis function and the resistance to chemotherapeutic injury.

Anemia ; drug therapy ; Animals ; Cells, Cultured ; Dose-Response Relationship, Drug ; Equidae ; blood ; Female ; Gelatin ; pharmacology ; Hematopoiesis ; drug effects ; Hematopoietic System ; drug effects ; Humans ; Male ; Mice ; Peptides ; pharmacology ; Serum Albumin ; pharmacology

The cancer-associated systemic inflammatory response is one of the critical indicators of tumor progression. Serum systemic inflammatory markers, such as neutrophil-lymphocyte ratio, lymphocyte-monocyte ratio, systemic inflammation score, Glasgow prognostic score, prognostic nutritional index, C-reactive protein-albumin ratio, lymphocyte-C-reactive protein ratio, platelet-lymphocyte ratio, are associated with the prognosis of colorectal cancer (CRC) . Further research of the prognostic value of inflammatory marks in CRC can provide help for the prognosis of CRC.

The fruit fly, Drosophila melanogaster, is able to discriminate visual landmarks and form visual long-term memory in a flight simulator. Studies focused on the molecular mechanism of long-term memory have shown that memory formation requires mRNA transcription and protein synthesis. However, little is known about the molecular mechanisms underlying the visual learning paradigm. The present study demonstrated that both spaced training procedure (STP) and consecutive training procedure (CTP) would induce long-term memory at 12 hour after training, and STP caused significantly higher 12-h memory scores compared with CTP. Label-free quantification of liquid chromatography-tandem mass spectrometry (LC-MS/MS) and microarray were utilized to analyze proteomic and transcriptomic differences between the STP and CTP groups. Proteomic analysis revealed 30 up-regulated and 27 down-regulated proteins; Transcriptomic analysis revealed 145 up-regulated and 129 down-regulated genes. Among them, five candidate genes were verified by quantitative PCR, which revealed results similar to microarray. These results provide insight into the molecular components influencing visual long-term memory and facilitate further studies on the roles of identified genes in memory formation.
Animals ; Conditioning (Psychology) ; physiology ; Drosophila Proteins ; genetics ; metabolism ; Drosophila melanogaster ; genetics ; metabolism ; physiology ; Flight, Animal ; physiology ; Gene Expression Profiling ; methods ; Memory ; physiology ; Oligonucleotide Array Sequence Analysis ; Proteomics ; methods ; Time Factors ; Vision, Ocular ; physiology

Objective To evaluate the relationship between the degree of brain injury during the perioperative period of liver transplantation and postoperative cognitive dysfunction (POCD).Methods Thirtythree patients,undergoing elective liver transplantation,were enrolled in this study.Before induction of anesthesia (T0),at 5 min before blocking the portal vein (T1),5 min after opening the portal vein (T2),5 min after opening the hepatic artery (T3),and at 24 h after surgery (T4),blood samples were collected from the central vein for determination of the serum concentrations of S1O0β protein and neuron-specific enolase (NSE) by enzyme-linked immunosorbent assay.Patients were divided into POCD group and control group (group C) according to whether POCD happened within 7 days after surgery.Results Compared with the baseline value at T0,the serum concentrations of S100β protein were significantly increased at T2 and T3,and the serum concentrations of NSE was increased at T3 (P＜0.05).There was no significant difference in serum concentrations of S100β and NSE at each time point between group POCD and group C (P＞0.05).Conclusion The degree of brain injury during the perioperative period of liver transplantation is not the dominant factor for the development of POCD in the patients.