Mfsd2a overexpression alleviates vascular dysfunction in diabetic retinopathy.

Diabetic retinopathy (DR) is one of the common complications in diabetic patients. Nowadays, VEGF pathway is subject to extensive research. However, about 27% of the patients have a poor visual outcome, with 50% still having edema after two years' treatment of diabetic macular edema (DME) with ranibizumab. Docosahexaenoic acid (DHA), the primary ω-3 long-chain polyunsaturated fatty acid (LC-PUFA), reduces abnormal neovascularization and alleviates neovascular eye diseases. A study reported that fish oil reduced the incidence of retinopathy of prematurity (ROP) by about 27.5% in preterm infants. Although ω-3 LC-PUFAs protects against pathological retinal neovascularization, the treatment effectiveness is low. It is interesting to investigate why DHA therapy fails in some patients. In human vitreous humor samples, we found that the ratio of DHA and DHA-derived metabolites to total fatty acids was higher in vitreous humor from DR patients than that from macular hole patients; however, the ratio of DHA metabolites to DHA and DHA-derived metabolites was lower in the diabetic vitreous humor. The expression of Mfsd2a, the LPC-DHA transporter, was reduced in the oxygen-induced retinopathy (OIR) model and streptozotocin (STZ) model. In vitro, Mfsd2a overexpression inhibited endothelial cell proliferation, migration and vesicular transcytosis. Moreover, Mfsd2a overexpression in combination with the DHA diet obviously reduced abnormal retinal neovascularization and vascular leakage, which is more effective than Mfsd2a overexpression alone. These results suggest that DHA therapy failure in some DR patients is linked to low expression of Mfsd2a, and the combination of Mfsd2a overexpression and DHA therapy may be an effective treatment.

Microsurgical Vitrectomy with Pars Plana Incision for the Removal of Posterior Segment Intraocular Foreign Bodies.

This study describes a pars plana incision surgical technique combined with 23 or 25-gauge vitrectomy in the management of intraocular foreign bodies (IOFBs) and to assess its anatomical and functional results. Sixteen patients with ocular trauma complicated with IOFB were enrolled in our study. The mean preoperative visual acuity was 2.01 ± 0.55 LogMAR, and the mean postoperative visual acuity at the final visit was improved to 0.91 ± 0.58 LogMAR (p < 0.001). Until the last follow-up, all IOFBs were successfully removed and anatomic success was obtained. Complications, such as endophthalmitis, silicone oil-dependent, and ocular hypotonia, were not observed. Microsurgical vitrectomy with modified pars plana incision is a safe and effective procedure in the treatment of retained IOFB, especially associated with transparent lens and posterior segment injury.

Relative activity of 15 bacterial strains against the larvae of Helicoverpa armigera, Spodoptera exigua, and Spodoptera litura (Lepidoptera: Noctuidae).

Crystal toxins produced by different strains of entomopathogenic Bacillus thuringiensis (Bt) have been characterized and widely applied as commercial biological pesticides owing to their excellent insecticidal properties. This study aimed to identify novel bacterial strains effective in controlling Spodoptera exigua Hübner, Helicoverpa armigera Hübner, and Spodoptera litura Fabricius. Fifteen culturable bacterial strains were isolated from 60 dead larvae (H. armigera and S. exigua) collected in the field. The biochemical characteristics and 16S rRNA sequences of these strains indicated that one strain (B7) was Lysinibacillus sp., 12 strains (B1, B3, B4, B5, B6, B8, P2, P3, P4, P5, P6, and DW) were Bt kurstaki, and P2-2 and B2 were Bacillus velezensis subsp. Laboratory bioassays indicated that strains B3, P6, B6, and P4 showed high toxicity to second-instar larvae of S. exigua, with LC50 values of 5.11, 6.74, 205.82, and 595.93 µg/ml, respectively; while the strains P5, B5, B6, and P6, were the most efficient against second-instar larvae of H. armigera with LC50 values of 725.82, 11,022.72, 1,282.90, 2,005.28, respectively, and strains DW, P3, P2, and B4 had high insecticidal activity against second-instar larvae of S. litura with LC50 values of 576.69, 1,660.96, 6,309.42, and 5,486.10 µg/ml, respectively. In conclusion, several Bt kurstaki strains with good toxicity potential were isolated and identified in this study. These strains are expected to be useful for biointensive integrated pest management programs to reduce the use of synthetic insecticides.

Feeding Spodoptera exigua larvae with gut-derived Escherichia sp. increases larval juvenile hormone levels inhibiting cannibalism.

Feed quality influences insect cannibalistic behavior and gut microbial communities. In the present study, Spodoptera exigua larvae were fed six different artificial diets, and one of these diets (Diet 3) delayed larval cannibalistic behavior and reduced the cannibalism ratio after ingestion. Diet 3-fed larvae had the highest gut bacterial load (1.396 ± 0.556 × 1014 bacteria/mg gut), whereas Diet 2-fed larvae had the lowest gut bacterial load (3.076 ± 1.368 × 1012 bacteria/mg gut). The gut bacterial composition and diversity of different diet-fed S. exigua larvae varied according to the 16S rRNA gene sequence analysis. Enterobacteriaceae was specific to the Diet 3-fed larval gut. Fifteen culturable bacterial isolates were obtained from the midgut of Diet 3-fed larvae. Of these, ten belonged to Escherichia sp. After administration with Diet 1- or 2-fed S. exigua larvae, two bacterial isolates (SePC-12 and -37) delayed cannibalistic behavior in both tested larval groups. Diet 2-fed larvae had the lowest Juvenile hormone (JH) concentration and were more aggressive against intraspecific predation. However, SePC-12 loading increased the JH hormone levels in Diet 2-fed larvae and inhibited their cannibalism. Bacteria in the larval midgut are involved in the stabilization of JH levels, thereby regulating host larval cannibalistic behavior.

[The effects of individualized therapeutic programs on chronic hepatitis C and the influential factors of virological response].

OBJECTIVE: To investigate the effect of individualized therapeutic programs with combination of interferon and ribavirin (RBV) in chronic hepatitis C (CHC) and study the influential factors of virological response rates. METHODS: A total of 139 patients with CHC were enrolled and given the intensive treatment doses of interferon and RBV according to their basic clinical condition. At the treatment of 0, 4, 12, 24 weeks, the end of treatment and 24 weeks after treatment stop, the serum HCV RNA was determined. Timely adjustment to dosage and time periods was made according to the virological response to treatment, and the predictive value of rapid virological response (RVR) and complete early virological response (cEVR) for sustained virological response (SVR) were analyzed. RESULTS: At the 4th week of treatment, the level of serum HCV RNA was monitored in 120 patients, and 84.2% (101/120) of patients obtained RVR; among them, 90.7% (88/97) obtained SVR. The virus load of patients obtained RVR at pretherapy was lower than that of patients didn't obtained RVR [(5.883 ± 1.246) lg copies/ml vs(6.502 ± 0.693) lg copies/ml, P = 0.034]. The RVR rate of initial treatment patients with PEG-IFNα-2a [87.8% (79/90)] was significantly higher than that of retreatment patients with PEG-IFNα-2a [65.0% (13/20)] (P = 0.031). At the 12th week of treatment, the level of serum HCV RNA was monitored in 132 patients, and 92.4% (122/132) of patients obtained cEVR; among them, 90.8% (108/119) obtained SVR. The SVR rate of patients obtained cEVR was significantly higher than that of patients didn't obtained cEVR (5/9) (P = 0.007). There was no significant difference between the cEVR rate of initial treatment patients [94.7% (90/95)] and retreatment patients [85% (17/20)] with PEG-IFNα-2a (P = 0.158). CONCLUSIONS: cEVR was predictor of SVR. Individualized therapy can increase the obtaining probability of RVR, cEVR and SVR. Adjusting drug dose timely and extending treatment period of HCV RNA-negative based on virological response to treatment are important in CHC individualized therapy.

[Detection of antibodies to cyclic citrullinated peptides and its clinical significance in patients with chronic hepatitis B virus infection].

OBJECTIVE: To detect the presence of antibodies to cyclic citrullinated peptides (anti-CCP) in patients with chronic hepatitis B virus (HBV) infection and evaluate its potential clinical significance. METHODS: Serum samples of 280 patients with chronic HBV infection and 40 healthy controls were collected from May 2011 to October 2011 and tested for anti-CCP and IgM-rheumatoid factor (RF). Anti-CCP was detected by ELISA and RF by immunonephelometry. All of 280 patients with chronic HBV infection were divided into 3 groups according to joint symptoms: asymptomatic group, HBV-associated arthropathy group and HBV concomitant RA group. Meanwhile, according to liver disease, they were divided into 3 groups: carrier group, chronic hepatitis group and cirrhosis group. RESULTS: The positive rates of anti-CCP and RF were 5.7% and 13.9% in patients with chronic HBV infection respectively. Anti-CCP was detected in 3 of 265 non-RA (1.1%) and 13 of 15 RA patients (86.7%). And RF were detected in 27 of 265 non-RA (10.2%) and 12 of 15 RA patients (80.0%). Twelve of 15 RA patients were positive for both anti-CCP and RF. The specificity of anti-CCP for RA was 98.9% in chronic HBV infection while the specificity of RF 89.8% (P < 0.01). Compared with the positive detection rates of anti-CCP and RF among liver disease subgroups, no significant difference existed between the subgroups. The levels of anti-CCP and RF in HBV concomitant RA group were statistically higher than those in asymptomatic group, HBV-associated arthropathy group and healthy controls (all P < 0.01). The level of RF in patients with HBV-associated arthropathy group was higher than that in asymptomatic group (U = 6017, P < 0.05). CONCLUSION: It is better to detect anti-CCP than RF to discriminate non-RA from concomitant RA in patients with chronic HBV infection.

Development of novel microsatellites for population genetic analysis of Phenacoccus solenopsis Tinsley (Hemipeta: Pseudoccoccidae) based on genomic analysis.

The cotton mealybug, Phenacoccus solenopsis Tinsley (Hemipeta: Pseudoccoccidae), is an aggressively invasive pest causing huge economic losses of crops around the world. In this study, we developed genome-wide microsatellites for population genetic analysis of P. solenopsis. We obtained a random genome of P. solenopsis with a size of 267.07 Mb and scaffold N50 of 14.12 Kb. In total 115,639 microsatellites were isolated from the genome, of which those with trinucleotide motifs were the most abundant. Forty-two polymorphic loci were selected for primer validation based on three populations. Allele numbers varied from 2 to 5 with an average value of 2.5 per locus, and allelic richness ranged from 1.00 to 4.48. The observed heterozygosity (H0) and expected heterozygosity (HE) ranged from 0.00 to 0.92 and 0.00 to 0.73, respectively. Population genetic structure analysis based on the developed markers revealed strong differentiation between three populations of P. solenopsis collected from its invasive range in China. The microsatellites developed in our study should provide efficient genetic markers for population level studies of P. solenopsis to reveal invasion history and patterns of dispersal.

[Matrix metalloproteinases inhibitor doxycycline inhibits TGF-beta 1-induced RPE cell migration].

OBJECTIVE: To investigate the effects of matrix metalloproteinases (MMP) inhibitor (Doxycycline) on retinal pigment epithelial (RPE) cell migration induced by transforming growth factor beta 1 (TGF-beta 1). METHODS: The third to sixth passage human RPE cells cultures were treated with TGF-beta 1 at different concentrations (0.01, 0.10, 1.00, 10.00 microg/L), and the conditioned media were collected after 36 h of TGF-beta 1 exposure. Gelatinase activities in conditioned media were analyzed by zymography. Migration assay was performed in a Boyden chamber with addition of different concentrations of Doxycycline. The number of migrated cells was counted under light microscopy. RESULTS: RPE cells migration were stimulated by TGF-beta 1 significantly, the number of migrated RPE cells increased about 27% compared with the control (P < 0.01). In the presence of Doxycycline, the RPE cells migration induced by TGF-beta 1 was inhibited and the number of migrated cells was reduced to 50% - 70% compared with control (P < 0.01). TGF-beta 1 also stimulated the secretion of MMP-2 in dose-dependent manner. CONCLUSIONS: This study indicates that Doxycycline can inhibit migration of RPE cells stimulated by TGF-beta 1. RPE migration induced by TGF-beta 1 may act partially through the stimulation of MMP production.

[Study on PBPs genes of penicillin-resistant Streptococcus pneumonia in Zhejiang province].

To elucidate alternations in gene/amino acid sequence of penicillin-binding proteins (PBPs) 1A, 2B, 2X from clinical isolates of penicillin-resistant Streptococcus pneumonia (PRSP) in Zhejiang Province, 26 strains of Streptococcus pneumonia were collected from November 2001 to January 2004. The antibiotics susceptibility of these strains was detected. PCR amplification and direct sequencing of PBP1A, 2B, 2X genes were performed. The sequence variations of PBP genes of the PRSPs in this region were studied by sequence BLAST analysis. It was shown that the main alternations of PBP1A were the four consecutive amino acid substitutions (Thr574Ala, Ser575Thr, Gln576Gly, Phe577Tyr) following the conservative motif KTG and the amino acid substitution Thr371Ser in the conservative motif STMK. The main alternation of PBP2B was Thr451Ala following the conservative motif SSN, and the main alternation of PBP2X was Thr338Ala in conservative motif STMK. The above mutation sites and drug resistant level were consistent to the data reported previously. Neither new gene mutation specific to these strains nor certain amino acid substitutions related to penicillin resistance reported was identified in the genes.

Association of IL10 -819C>T and -592C>A Polymorphisms with Non-Hodgkin Lymphoma Susceptibility: Evidence from Published Studies.

Numerous studies have investigated the association of IL10 -819C>T and -592C>A polymorphisms with non-Hodgkin lymphoma (NHL) susceptibility, and yet reported conflicting results. With this in mind, we performed the current meta-analysis with an aim to verify actual causative variants underlying lymphomagenesis. Pooled odds ratios (ORs) and 95% confidence intervals (CIs) were calculated to evaluate the strength of the associations. Moreover, to explore the biological function of these polymorphisms, we also performed genotype-based mRNA expression analysis using online database derived from 270 subjects within three ethnicities. The final analysis included 11 studies with a total of 5859 NHL cases and 6893 controls for the IL10 -819C>T polymorphism, and 11 studies with 6277 cases and 7350 controls for the IL10 -592C>A polymorphism. No significant association was observed for these two polymorphisms in either the overall analysis or the stratification analyses by ethnicity and source of controls. Nevertheless, stratification analyses demonstrated a significant decreased risk associated with the IL10 -819C>T polymorphism (homozygous: OR=0.81, 95% CI=0.66-0.99, and recessive model: OR=0.80, 95%CI=0.65-0.98) and IL10 -592C>A polymorphism (homozygous: OR=0.80, 95% CI=0.66-0.99, and recessive model: OR=0.80, 95%CI=0.66-0.97) among patients with diffuse large B-cell lymphoma (DLBCL). Despite some limitations, this meta-analysis indicates that polymorphisms in IL10 gene may contribute to DLBCL susceptibility.

[Influence of fruit size of Camellia meiocarpa on growth of oil tea weevil, Curculio chinensis (Coleoptera: Curculionidae)].

The relationship between mature larval mass of oil tea weevil (Curculio chinensis) and fruit volume of its host plant oil tea (Camellia meiocarpa) was fitted with Logistic equation in order to understand the restriction of host fruit size on large larval growth and development of the weevil. The results showed that the larval mass increased with the increasing host fruit volume, which was in good conformity with the Logistic model. The weevil larval growth followed the principle of diminishing marginal utility, and it could be divided into two periods, the fast-growing period (<3.216 cm3, one larva per fruit; <4.747 cm3, two larvae per fruit ) and the asymptotic growing period (>3.216 cm3, one larva per fruit; >4.747 cm3, two larvae per fruit). The minimum fruit size threshold was 1500 cm3 for one larva per fruit, and 2.539 cm3 for two larvae per fruit. The temporal pattern that the mature larvae exited from their host fruits was established, the number of larvae escaping from their host fruits decreased daily after the fruit collection, and the larval escaping peak largely appeared from 6:00 to 10:00 AM with 43.9% of total escaping number, and especially from 7:00 to 8:00 AM with 21.1% of total escaping number. The bigger the larvae, the earlier exited from their host fruits. The restriction of fruit size on growth and development of oil tea weevil was observed, and it should be a behavioral adaptation strategy to increase the offspring' s fitness for the parental weevil adults to oviposit on the bigger fruits.

[Detection of BK virus infection in renal transplant recipients and clinical application].

OBJECTIVE: To study the detection methods of BK virus infection in kidney transplant recipients, and to explore the clinical application. METHODS: 132 cases of renal transplant recipients were undertaken BK virus detection including presence of decoy cells in urinary sediment, urine and serum BKV-DNA to demonstrate the BK virus replication. RESULT: Among 132 cases of renal transplant recipients, urinary decoy cell was found in 37 (28.0%) patients and the median time was 12 months after surgery. 32 (24.2%) patients were diagnosed as BK viruria at a median of 11 months after surgery, and 16 (12.1%) recipients were diagnosed as BK viremia at a median of 15 months after surgery, 5 patients with BK viruria were diagnosed as BK virus associated nephropathy according to allograft biopsy. CONCLUSION: To make early diagnosis of BK virus infection, detection of urine decoy cells and BKV-DNA in urine and plasma sample is important,which provides an important basis for the prevention of BK virus associated nephropathy.

[Clinical features and gene mutation analysis in Machado-Joseph disease of spinocerebellar ataxia type 3 in littoral of Zhejiang].

OBJECTIVE: To study the clinical features and gene mutation analysis in Machado-Joseph disease of spinocerebellar ataxia type 3 in littoral of Zhejiang. METHODS: Clinical manifestation and brain MRI data 18 patients with SCA in family were analyzed. The gene mutations of 18 patients and 10 family numbers without abnormal presentation, and 12 healthy persons of controls. RESULTS: The gene mutations of 18 patients is SCA3/MJD, and 2 asymptomatic SCA3/MJD had been detected in SCA family. Normal alleles of SCA3/MJD have CAG repeats ranging from 14 to 27, patients from 67 to 82, asymptomatic and carrier SCA3/MJD from 28 to 45. The main features of 18 patients included gait ataxia, ambiguity in speech and action clumsiness. Brain MRI showed remarkable atrophy on cerebellum and brain stem. CONCLUSION: CAG expansions were related to SCA3/MJD. The clinical manifestations are ataxia and dysarthria. The detection of repeated times CAG can provide an effective way for the genetic and asymptomatic diagnosis.

[Expression of recombinant VP2 gene in insect sf9 cells and screening of clinical specimens].

OBJECTIVE: To clone and express VP, gene from HBoV, and the expressed VP, protein was as the antigen in order to detect serum from children in Wenling area with lower respiratory tract infections. METHODS: The VP, gene was recombined with the genome of Baculovirus, which infected the insect cell. The fusion protein with HA tag was applied to confirm the specificity of expressed protein. Furthermore, the recombinant protein was observed using electron microscopy. The 176 serum from children in Wenling area with lower respiratory tract infections was screened using Western blot. RESULTS: The expressed VP2 protein was more than 60% in total proteins from insect cell, and MWt about 60 x 10(3). The virus-like particle (VLP) was observed using electron microscopy, and size about 20 nm. The 176 serum from children in wenling area with lower respiratory tract infections was screened using Western blot. The HBoV positive rate was 2.28% (4/176). CONCLUSION: The VP2 protein from human bocavirus was expressed in insect cell successfully. Through HA tag the VP2 protein was specific, and then the assay using SDS-PAGE with Western blot could detect and screen the antibody in serum from children with lower respiratory tract infections rapidly and accurately.

[Isolation and cell culture of human bocavirus (HBoV) by human bronchial epithelial cell lines].

OBJECTIVE: To investigate pave a way for studying pathogenicty of HBoV. METHODS: Isolation and cell culture of HBoV by human bronchial epithelial cell line, which was founded in our laboratory. The morphology of the virus were primarily studied with a transmission electron microscope. In addition, transcript mRNA was detected in human bronchial epithelial cells, which was passaged and infected within HBoV, using the reverse-transcription polymerase chain reaction (RT-PCR). The amplified products nucleotide sequence of HBoV were sequencing and sequence analysis. RESULTS: Cytopathic effect (CPE) was observed after the aseptic residue of filtration of 2 case sputum specimens with HBoV, which was inoculated to the human bronchial epithelial cell line. The virus particles were observed in the cytoplasm, which were hexagonal or spherical in shape and 18-26 nm in diameter,bulk was 20 nm. cDNA amplicon obtained 295 bp fragment results of electrophoresis bands as same as NS1 region of the conserved matrix gene of publish sequence of HboV. PCR products nucleotide sequence of HboV were compared with corresponding HboV GeneBank sequences. The comparison/alignment and construction of phylogenetic trees also point to an affiliation of the parvovirus to the species HBoV. CONCLUSION: Isolation and identification of HBoV could be done in the human bronchial epithelial cell, and we found some characterizing CPE in the human bronchial epithelial cell after HBoV infection. The above studies pave a way for studying pathogenicty of human bocavirus.

[Discovery and identification of WU polyomavirus in children from Zhejing region].

WU polyomavirus, which was firstly discovered in 2007, is a new human polyomavirus belonging to Polyomaviridae and containing circular double-stranded genomic DNA. In this study, the 278 clinical sputum specimens from children under 5 years old were collected from Wenzhou Medical College affiliated Wenling First Hospital, Zhejiang Province. Based on identification assay of WU polyomavirus previously reported, a WU polyomavirus was identified from clinical samples successfully, the positive rate was 0.4%. The sequences of PCR products were identical to that of VP2 gene and large T antigen gene derived from WU polyomavirus reported. The above results strongly suggested that the WU polyomavirus isolated was firstly found in Chinese children with acute lower respiratory tract infections. This study provides a firm basis for further research of WU polyomavirus.

[Characterization of the cytopathic effect in human bronchial epithelial cell after Human Bocavirus Infection (HBoV)].

OBJECTIVE: In this study, human bronchial epithelial cells were inoculated with positive sputum specimens of HBoV. After four days' infection, cytopathic effects (CPE) were observed by inverted microscopy. These viruses all cause typical cell damages such as rounded and shrivelled, fusion and fallout. These damages got quick following increased future degenerations. The other assay result of CPE within the infected cells were observed by inverted microscopy, have typical "owl's eye" plaque and above 90 percent hemadsorption within the infected cells by erythrocytes for hemadsorption technique. The typical fluorescence lump of nucleus within the infected cells was found by indirect immunofluorescence technique. CONCLUSION: Isolation and identification of HBoV could be done in the human bronchial epithelial cell, and we found some characterizing CPE in the human bronchial epithelial cell after HBoV infection. The above studies pave a way for studying pathogenicity of human bocavirus.

[Clinical application of microarray technique to quantify the lamivudine-resistant gene mutant of hepatitis B virus].

OBJECTIVE: To evaluate clinical applicability of a novel technique that can quantify the lamivudine-resistant mutants of hepatitis B virus (HBV) in the serum of patients utilizing gene microarray technology. METHODS: The oligonucleotide microarray was designed to detect 3 important mutational positions. Fifty-one patients who were receiving lamivudine therapy were selected as subjects. The oligonucleotide microarray and traditional sequencing were applied to detect the lamivudine resistant mutation, the monitoring lasted for 24 months. Then the clinical result was analyzed and the obtained data were compared between the two methods. RESULTS: Lamivudine resistant mutation was detected in 39 percent of the patients during the 2 years period. The results of the oligonucleotide microarray technique was consistent to the results of traditional sequencing in accuracy and the miroarray was more sensitive in detection of the mixed infection. CONCLUSION: Application of the oligonucleotide microarray for quantitative detection of lamivudine-resistant mutation of HBV is feasible.

[Clinical prospective study on maternal-fetal transmission of human bocavirus].

OBJECTIVE: To investigate maternal-fetal transmission at human bocavirus (HBoV). METHODS: IgG antibody to HBoV in serum samples of 316 mothers were determined with ELISA and HBoV DNA was determined with real time PCR in the sera of the mothers and their infants. RESULTS: HBoV-IgG was positive in 40.20 percent (127/316) of the mothers, while it was positive in 29.43 percent (93/316) of the cord blood specimens of the infants. The difference between the two groups was significant (X2=8.12, P less than 0.005); 93 samples of both the mothers and the infants were positive for HBoV-IgG. CONCLUSION: HBoV-IgG can cross the placenta to the fetuses through placenta. Further study is needed to answer the question whether vertical maternal-fetal transmission occurs.