OBJECTIVETo investigate the function of POLH(polymerase eta) through establishment of the POLH gene-blocked cell line FL-POLH(-).

METHODSA mammalian expression vector expressing antisense POLH gene fragment pMAMneo-amp-POLHA (-) was constructed by cloning the 1473 - 2131 fragment of POLH gene into the mammalian expression vector pMAMneo-amp(-) in antisense orientation. The FL cells were transfected with this antisense RNA expressing vector and selected by G418. The mutation assay was conducted using the shuttle plasmid pZ189.

RESULTThe spontaneous mutation frequency of SupF tRNA gene in the plasmid replicated in the FL-POLH(-) was 13.5 x 10(-4), while it was 4.9x10(-4) and 3.7x10(-4) in the control cells FL and FL-M, respectively. The nontargeted mutation frequency of SupF tRNA gene decreased in the plasmid replicated in these cell lines pretreated with MNNG.

CONCLUSIONPOLH plays an important role in maintenance of genetic stability and genesis of nontargeted mutation.

Antisense Elements (Genetics) ; pharmacology ; Cell Line ; DNA-Directed DNA Polymerase ; genetics ; physiology ; Methylnitronitrosoguanidine ; toxicity ; Mutagenesis

With the development of society, the new requirements for visual health are put forward. In the past, visual acuity and contrast sensitivity were used to evaluate the visual function of human eye, but it is subjective and limited, not really reflecting human eye's visual function. In order to pursue a more sensitive and accurate objective evaluation method, experts at home and abroad make some exploratory research on visual quality. The study confirmed that the scattering is an independent factor affecting the visual quality of human eye, it can make a more scientific evaluation of visual function.This article summarizes the influence of scattering for the visual quality of human eye, according to related research at home and abroad in recent years.

Objective:To compare the postoperative adjacent segment degeneration (ASD) between the microscopically anterior cervical discectomy with fusion (ACDF) and anterior cervical corpectomy with fusion (ACCF) in the treatment of cervical spondylotic myelopathy and its influencing factors.Methods:Fifty patients with cervical spondylotic myelopathy treatment in the Qinzhou Second People′s Hospital from July 2018 to July 2020 were selected, they were divided into two groups, 25 patients performed ACDF (ACDF group), and 25 patients performed ACCF (ACCF group). The perioperative period, efficacy and incidence of ASD were compared between the two groups, and the influencing factors of ASD were analyzed.Results:The intraoperative blood loss, operation time, length of hospital stay and postoperative drainage in ACCF group were higher than those in ACDF group: (58.34 ± 8.61) ml vs. (46.77 ± 7.24) ml, (99.57 ± 10.72) min vs. (86.14 ± 9.64) min, (8.97 ± 1.43) d vs. (7.56 ± 1.24) d, (17.92 ± 2.95) ml vs. (14.28 ± 2.66) ml, there were statistical differences ( P<0.05). The postoperative Japanese Orthopaedic Association (JOA) scores and Neck Disability Index (NDI) scores in the two groups were improved significantly ( P<0.05), but the scores of JOA and NDI in the two groups had no significant differences ( P>0.05). The incidence of ASD in the two groups had no significant differences ( P>0.05). The Cox univariate analysis showed that age >59 years, intervertebral disc degeneration, number of fusion segments >2, osteoporosis and postoperative ASD were risk factors for ASD( P<0.05). Conclusions:The effect of microscopically ACDF is similar to that of ACCF in the treatment of cervical spondylotic myelopathy, but ACDF has the advantages of less trauma and quick recovery. The risk of postoperative ASD should be vigilant for patients with age >59 years old, intervertebral disc degeneration, number of fusion segments >2 or osteoporosis.

Objective:To investigate the role of TcpC in uropathogenic Escherichia coli (UPEC)-induced cystitis in mice and to preliminarily analyze the pathogenic mechanism. Methods:C57BL/6 mice were injected with 10 9 CFU wild-type UPEC CFT073 (CFT073 wt) or tcpc gene-deleted mutant (CFT073 Δ tcpc) from urethra into bladder to construct the mouse model of cystitis. The mice were sacrificed 3 d after infection and the bladders were taken to observe the gross pathological changes. Histopathological changes in bladder tissues were observed after HE staining. Immunohistochemistry was used to detect TcpC in bladder tissues. Bacterial loads in urine samples of UPEC-infected mice were counted by tenfold dilution method, and the presence of tcpc gene in the genomic DNA of bacteria from the bladder and urine samples of CFT073 wt-infected mice was measured by PCR. Real-time quantitative RT-PCR (qRT-PCR) and Western blot were performed to detect the expression of TcpC at mRNA and protein levels in macrophages after CFT073 wt infection. The influence of UPEC strains on the activation of NF-κB signaling pathway in macrophages were determined by Western blot. The levels of proinflammatory factors and the bacterial and cell activity after infecting macrophages with UPEC strains were detected by ELISA, laser confocal microscope and fluorescence microscope, respectively. Results:Compared with the mice with CFT073 Δ tcpc infection, CFT073 wt-infected mice had significantly enlarged bladder and severe neutrophil infiltration and abundant TcpC in bladder tissues. The number of bacteria in the urine of CFT073 wt-infected mice was significantly greater than that of the CFT073 Δ tcpc group. PCR results showed that the bacteria in bladder or urine were CFT073 wt. The expression of TcpC at both mRNA and protein levels in macrophages increased significantly after CFT073 wt infection. Moreover, in CFT073 wt-infected macrophages, the expression of IκBα was promoted and the phosphorylation of p65 and the production of proinflammatory factors were suppressed. TcpC was instrumental in the survival and invasion of CFT073 wt in macrophages. Conclusions:TcpC expression increased significantly in mice with CFT073 wt-induced cystitis. TcpC inhibited the activation of NF-κB signaling pathway and the production of proinflammatory factors in macrophages to improve the survival rate of CFT073 wt, which was closely related to the pathogenesis and immune evasion of UPEC.

OBJECTIVE: To study the clinical screening and genetic diagnosis of children suspected of Prader-Willi syndrome (PWS), as well as the differences in the scores of clinical diagnostic criteria among the children with a confirmed diagnosis of PWS. METHODS: A total of 94 children suspected of PWS who were admitted from July 2016 to December 2018 were enrolled as subjects. Methylation-specific multiplex ligation-dependent probe amplification (MS-MLPA) was performed to confirm the diagnosis. For the children with a confirmed diagnosis of PWS, the scores of clinical diagnostic criteria were determined, and the perinatal characteristics were analyzed. RESULTS: A total of 11 children with PWS were confirmed by MS-MLPA, with a detection rate of 12%, among whom there were 7 boys and 4 girls, with a median age of 3 years and 4 months (range 25 days to 6 years and 8 months) at the time of confirmed diagnosis. Among the 11 children with PWS, only 5 children (45%) met the criteria for clinical diagnosis. The main perinatal characteristics of the children with PWS were decreased fetal movement (9 cases, 82%), cesarean section birth (11 cases, 100%), hypotonia (11 cases, 100%), feeding difficulties (11 cases, 100%), and weak crying (11 cases, 100%). CONCLUSIONS Gene testing should be performed as early as possible for children suspected of PWS by clinical screening. PWS may be missed if only based on the scores of clinical diagnostic criteria.
Cesarean Section ; Child ; Child, Preschool ; Female ; Humans ; Infant ; Infant, Newborn ; Male ; Methylation ; Muscle Hypotonia ; Prader-Willi Syndrome ; Pregnancy

This article reports two cases of childhood-onset nemaline myopathy diagnosed by muscle pathology and genetic diagnosis. The two patients had onset in early childhood, with muscle weakness as the first manifestation, as well as long disease duration and slow progression. Gomori staining and hematoxylin-eosin staining showed red-stained rods in the sarcoplasmic cytoplasm and sarcolemma under a light microscope. Electron microscopy showed that the dense nemaline rods were located under the muscle fiber sarcolemma and parallel to the long axis of the muscle fibers, and some muscle fiber myofilaments were dissolved and necrotic. Gene testing found that one of the two patients had heterozygous mutation (c.1013A>C) in the ACTA1 gene, and the other had compound heterozygous mutation (c.18676C>T and c.9812C>A) in the NEB gene. The two mutations were more common in nemaline myopathy. Nemaline myopathy is a recessive or dominant inheritance myopathy, in which the nemaline rod in the cytoplasm of myocytes is a characteristic muscle pathological change. Pathological and genetic diagnosis is the gold standard for diagnosis of nemaline myopathy.

This study was aimed to explore whether the conditioned culture medium of human umbilical cord-derived mesenchymal stem cells (hUC-MSC) has supportive effects on hematopoiesis in vitro. hUC-MSC were cultured in 75 cm(2) culture flasks at a concentration of 2×10(6) cells per flask. After 48 h, the conditioned culture medium was harvested. CD34(+) cells were isolated with the human cord blood CD34 positive selection kit. The CD34(+) cells were plated in three different culture systems: the culture supernatant from hUC-MSC added into incomplete methylcellulose without recombinant human cytokines as conditioned culture medium; the complete methylcellulose medium with recombinant human cytokines as positive control medium; incomplete methylcellulose adding DMEM/F12 with 10% FBS instead of conditioned culture medium as the negative control medium. After 14 days of culture, colonies containing ≥ 50 cells were scored and types of colonies were classified under inverted microscope. The immunophenotypes of cells which were collected from the colonies were detected by flow cytometry. The results showed that conditioned culture medium of hUC-MSC supported the differentiation of CD34(+) cells into CFU-G (47.67 ± 0.58), CFU-GM (48.67 ± 4.73) and CFU-M (3.00 ± 2.00) in vitro, while the CFU-E, BFU-E or CFU-GEMM were absent. Comparatively, in the positive control medium all kinds of CFU were observed. Interestingly, the percentage of CD45(+)cells of CFU in conditioned culture medium (97.43 ± 2.15)% was more than CD45(+)cells in positive control medium (39.69 ± 0.96)% (P < 0.05). It is concluded that the conditioned culture medium of hUC-MSC has been confirmed to have ability to support hematopoiesis separately in vitro. Besides, it enhances the differentiation of CD34(+) cells into myeloid cells except cells of erythroid lineage.
Antigens, CD34 ; Cell Differentiation ; Cells, Cultured ; Culture Media, Conditioned ; Fetal Blood ; cytology ; Hematopoiesis ; Humans ; Mesenchymal Stromal Cells ; cytology ; Umbilical Cord ; cytology

VisualiZation detection methods are used for determination of the concentration of unknown target by comparing the color change in the intensity or type of reaction solution by naked eye.VisualiZation detection method has some advantages such as simple and rapid operation, low detection cost, fast reaction speed, and detecting target concentration by means of naked eye.Gold nanomaterials are widely used in the construction of visual biosensors due to its unique optical properties.For example, when changing the distance or morphology of the particles, the plasmon resonance absorption peak of local surface will change accordingly.Herein, we reviewed the application of gold nanomaterials in visualiZation biosensors for the detection of target molecules, summed up the main problems of AuNP colormertic methods in the determination of actual samples, and provided an outlook of the future of gold nanoparticles-based biosensor in application development.

OBJECTIVETo investigate the association of -1131T>C and c.553G>T polymorphisms and their haplotypes in apolipoprotein A5(ApoA5) gene with cereberovascular disease in Chinese.

METHODSUsing polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP), we analyzed two ApoA5 genetic variants in 272 patients with cerebral infarction (CI) and 316 control individuals respectively. The levels of serum lipid profiles were measured with biochemical methodsìand the other clinical characters were obtained by case file investigation.

RESULTSThe odds ratio (OR) for CI in -1131CC genotype carriers was 2.10 (95%CI 1.01-4.37). The distribution of T-T and T-G haplotypes had obvious differences between CI patients and control individuals. The OR for CI in C-G and T-G haplotype carriers were 1.34 and 0.71(95% CI 1.02-1.76 and 0.55-0.92) respectively, compared with the others. Furthermore, the major haplotypes had significant differences of serum TG(P< 0.05).

CONCLUSIONThe ApoA5 -1131T>C polymorphism may be associated with an increased risk of CI in the Chinese population, but the influence of blood lipids can not be ignored.

Aged ; Apolipoproteins A ; genetics ; Carotid Artery Diseases ; complications ; genetics ; Cerebral Infarction ; complications ; genetics ; Female ; Genetic Predisposition to Disease ; genetics ; Haplotypes ; genetics ; Humans ; Male ; Middle Aged ; Polymerase Chain Reaction ; Polymorphism, Genetic ; genetics ; Polymorphism, Restriction Fragment Length

OBJECTIVETo analyze the relationship between orbital fat volume and the progression and prognosis of thyroid- associated ophthalmopathy (TAO) and determine the optimal treatment timing for TAO.

METHODSThe clinical data were collected from 35 patients (70 orbits) with a definite diagnosis of TAO between January, 2016 and December, 2016. The correlation between orbital fat volume and the clinical parameters was evaluated. We also analyzed the correlation of the signal intensity ratio (SIR) of the extraocular muscles with the clinical parameters. The orbital fat volume was compared between patients with TAO and 12 control subjects.

RESULTSThe orbital fat volume was significantly correlated with the duration of TAO (r=0.480, P<0.01), but showed no significant difference between patients with a disease course within 6 months and those with a disease course of 6 to 12 months (P=0.084). The patients with a disease course beyond 12 months had a significantly greater orbital fat volume than those with a disease course of 6 months (P<0.01) or 6 to 12 months (P<0.05). The orbital fat volume was correlated with the degree of proptosis (r=0.622, P<0.01), and an increase of exophthalmos by 1 mm was associated with a total orbital volume increment of 0.88 mL. The clinical activity score was correlated with the SIR of the extraorbital muscles (r=0.536, P<0.01) and levels of anti-thyroid-stimulating hormone receptor antibody (r=0.416,P<0.01). The orbital fat volume was significantly greater in TAO patients than in the healthy individuals (P<0.01).

CONCLUSIONIn patients with TAO, the peak increase of orbital fat volume occurs one year after the disease onset. Measurement of the orbital fat volume combined with SIR of the extraorbital muscles can serve as an indicator for determining the optimal timing for intervention of TAO and helps in the evaluation of prognosis of the patients.