Objective:To investigate the factors influencing phytohemagglutinin (PHA) response in the detection of Mycobacterium tuberculosis infection by gamma interferon release assay (IGRA). Methods:A retrospective case-control study was conducted on 360 hospitalized patients who received IGRA in West China Hospital of Sichuan University from January 2019 to December 2021. According to PHA response (IFN-γ level), they were divided into three groups: negative mitogen response group (IFN-γ<2 pg/ml), weak positive mitogen response group (IFN-γ: 2-100 pg/ml), and normal mitogen response group (IFN-γ>400 pg/ml).Results:Immune diseases were independently associated with negative (OR=0.34, 95%CI: 0.17-0.72, P=0.004) and weak positive mitogen responses (OR=0.29, 95%CI: 0.16-0.55, P<0.001). Infections caused by pathogens other than Mycobacterium tuberculosis was independently associated with negative mitogen response (OR=0.266, 95%CI: 0.09-0.83, P=0.023), while immunodeficiency was independently associated with weak positive mitogen response (OR=0.280, 95%CI: 0.12-0.63, P=0.002). Mitogen response was significantly correlated with the levels of albumin and hemoglobin in serum and the counts of neutrophils and lymphocytes ( P<0.001). Conclusions:Immune diseases and immunodeficiency can affect mitogen response. Therefore, clinicians should give attention to mitogen response in the interpretation of IGRA test results to prevent misdiagnosis and underdiagnosis. Besides, to a certain extent, mitogen response can reflect the infection status of hospitalized patients.

Objective To explore the role of zinc finger protein 36 (ZFP36) in regulating osteogenic differentiation of bone marrow-derived mesenchymal stem cells (BMSCs) and preosteoblasts. Methods ZFP36 expression was observed in primary mouse BMSCs and mouse preosteoblasts (MC3T3-E1 cells) during induced osteogenic differentiation. Zfp36-deficient cell models were constructed in the two cells using RNA interference technique and the changes in differentiation capacities of the transfected cells into osteoblasts were observed. Transcriptome sequencing was used to investigate the potential mechanisms of ZFP36 for regulating osteoblast differentiation of the two cells. U0126, a ERK/MAPK signal suppressor, was used to verify the regulatory mechanism of Zfp36 in osteogenic differentiation of Zfp36-deficient cells. Results During the 14-day induction of osteogenic differentiation, both mouse BMSCs and MC3T3-E1 cells exhibited increased expression of ZFP36, and its mRNA expression reached the peak level on Day 7 (P<0.0001). The Zfp36-deficient cell models showed reduced intensity of alkaline phosphatase (ALP) staining and alizarin red staining with significantly lowered expressions of the osteogenic marker genes including Alpl, Sp7, Bglap and Ibsp (P<0.01). Transcriptome sequencing verified the reduction of bone mineralization-related gene expressions in Zfp36-deficient cells and indicated the involvement of ERK signaling in the potential regulatory mechanism of Zfp36. Immunoblotting showed that pERK protein expression increased significantly in Zfp36-deficient cells compared with the control cells. In Zfp36-deficient MC3T3-E1 cells, inhibition of activated ERK/MAPK signaling with U0126 resulted in obviously enhanced ALP staining and significantly increased expressions of osteoblast differentiation markers Runx2 and Bglap (P<0.05). Conclusions ZFP36 is involved in the regulation of osteoblast differentiation of mouse BMSCs and preosteoblasts, and ZFP36 deficiency causes inhibition of osteoblast differentiation of the cells by activating the ERK/MAPK signaling pathway.

Objective To explore the role of zinc finger protein 36 (ZFP36) in regulating osteogenic differentiation of bone marrow-derived mesenchymal stem cells (BMSCs) and preosteoblasts. Methods ZFP36 expression was observed in primary mouse BMSCs and mouse preosteoblasts (MC3T3-E1 cells) during induced osteogenic differentiation. Zfp36-deficient cell models were constructed in the two cells using RNA interference technique and the changes in differentiation capacities of the transfected cells into osteoblasts were observed. Transcriptome sequencing was used to investigate the potential mechanisms of ZFP36 for regulating osteoblast differentiation of the two cells. U0126, a ERK/MAPK signal suppressor, was used to verify the regulatory mechanism of Zfp36 in osteogenic differentiation of Zfp36-deficient cells. Results During the 14-day induction of osteogenic differentiation, both mouse BMSCs and MC3T3-E1 cells exhibited increased expression of ZFP36, and its mRNA expression reached the peak level on Day 7 (P<0.0001). The Zfp36-deficient cell models showed reduced intensity of alkaline phosphatase (ALP) staining and alizarin red staining with significantly lowered expressions of the osteogenic marker genes including Alpl, Sp7, Bglap and Ibsp (P<0.01). Transcriptome sequencing verified the reduction of bone mineralization-related gene expressions in Zfp36-deficient cells and indicated the involvement of ERK signaling in the potential regulatory mechanism of Zfp36. Immunoblotting showed that pERK protein expression increased significantly in Zfp36-deficient cells compared with the control cells. In Zfp36-deficient MC3T3-E1 cells, inhibition of activated ERK/MAPK signaling with U0126 resulted in obviously enhanced ALP staining and significantly increased expressions of osteoblast differentiation markers Runx2 and Bglap (P<0.05). Conclusions ZFP36 is involved in the regulation of osteoblast differentiation of mouse BMSCs and preosteoblasts, and ZFP36 deficiency causes inhibition of osteoblast differentiation of the cells by activating the ERK/MAPK signaling pathway.

Objective To explore the potential regulatory mechanism of resistance exercise in senescence accelerated-prone 8 mice(SAMP8)by evaluating the effects of exercise on the expression of long non-coding RNA(lncRNA)and mRNA in quadriceps muscles by RNA-sequencing(RNA-seq)technology.Methods Twenty-eight-week-old male SAMP8 mice were divided into a model group(M group)and resistance-exercise group(R group)(n=6 mice per group).Another eight normally aging SAMR1 mice of the same age were used as the control group(C group).Mice in R group received 8 weeks of increasing weight climbing exercise training.Relative grip strength was measured every week and the rotarod test was performed every 2 weeks.Histological changes in the right quadriceps femoris were observed by hematoxylin-eosin staining and the left quadriceps was used for RNA-seq.Differentially expressed lncRNA and mRNA were screened and analyzed for enrichment by Gene Ontology(GO)and Kyoto Encyclopedia of Genes and Genomes(KEGG)analyses.Finally,key differentially expressed genes were analyzed by quantitative reverse transcription-polymerase chain reaction to verify the accuracy of the RNA-seq result.Results(1)Relative grip strength and rotarod test time were significantly decreased in M group compared with C group(P＜0.01),but were significantly increased after 8 weeks of Rgroup compared with M group(P＜0.01).(2)The cross-sectional area of the muscle fibers was significantly lower in M group compared with C group,as shown by HE staining(P＜0.01),while the cross-sectional area of the muscle fibers was significantly increased in the R group compared with M group(P＜0.01).(3)Differential expression analysis identified 182 upregulated and 218 downregulated lncRNA,and 454 upregulated and 289 downregulated mRNA between M group and R group.The KEGG pathways of lncRNA target genes that were differentially expressed between M group and R group were significantly enriched in intestinal immune network for IgA production,nuclear factor-kappa B signaling pathway,and inflammatory bowel disease.Conclusions(1)This study demonstrated that resistance exercise can improve skeletal muscle function in SAMP8 mice with sarcopenia.We identified lncRNA and mRNA that were differentially expressed as a result of resistance exercise,and which might be potential targets of sarcopenia therapy.(2)Furthermore,analyzing the biological functions of the target genes of the differentially expressed lncRNA and mRNA may further our understanding of the mechanism of resistance exercise for improving sarcopenia.

Objective:To analyze the epidemiological characteristics of respiratory virus infections in Sichuan from 2017 to 2023, and investigate the changes in detection rates of various viruses in different periods and age groups.Methods:The study included 41 516 patients with acute respiratory infections. Clinical laboratory results of these patients were retrospectively collected to calculate the detection rates of various viruses. The epidemiological characteristics of respiratory virus infections before, during, and after the COVID-19 pandemic, the prevalence of mixed infections, and the detection rates in different seasons and age groups were analyzed.Results:The overall detection rate of respiratory viruses was 23.35% (9 696/41 516), with influenza A virus (8.47%, 3 515/41 516), rhinovirus (7.13%, 2 961/41 516), and influenza B virus (2.04%, 846/41 516) being the most prevalent. The highest detection rate was observed in children aged 0-14 years (48.62%, 1 429/2 939), while the detection rates in young, middle-aged, and elderly individuals were similar [21.00% (1 482/7 057), 22.06% (3 828/17 350), 20.33% (2 685/13 205)]. The detection rate of influenza A virus dropped to 1.35% (253/18 717) during the COVID-19 pandemic, and rebounded to 13.05% (1 560/11 953) after the pandemic. A similar pattern was observed in other viruses such as human metapneumovirus, influenza B virus, coronavirus, and respiratory syncytial virus, with significant reductions during the COVID-19 pandemic and subsequent increases that even exceeded pre-pandemic levels. The detection rates of adenovirus, bocavirus, and rhinovirus gradually decreased over time, while the prevalence of parainfluenza virus remained stable, indicating a less significant impact of COVID-19 on these viruses. Higher detection rates were observed in autumn (28.90%, 2 324/8 041) and winter (26.52%, 4 093/15 433) as compared to that in spring (17.61%, 1 848/10 494) and summer (18.96%, 1 431/7 548), suggesting significant seasonal impacts on virus detection rates ( P<0.001). This study identified 8 666 cases of single-virus infections, 948 double infections, 77 triple infections, and five quadruple infections. Co-infection of bocavirus and rhinovirus was the most common type of double infections (17.41%, 165/948), and 59.60% (565/948) of double infections involved rhinovirus. Conclusions:The overall detection rate of respiratory viruses is higher in autumn and winter than in spring and summer. The prevalence of some respiratory viruses such as influenza viruses decreases during the COVID-19 pandemic. People of different ages can develop multiple infections with rhinovirus being the most common in double infections.

Objective: To explore the visuospatial memory characteristics of school-age children with exotropia and to analyze associated factors. Methods: Based on a case-control study,45 exotropia children aged 8-12 years and 45 normal control children were recruited from 2017-2019. The "tapping test" was used to evaluate the visuospatial short-term and working memory of children. Results: There was no significant differences in the scores of visuospatial short-term memory between the exotropia group and the control group [(7.64±1.69)(8.00±1.66)，t=-1.00，P=0.32)]. The scores of visuospatial working memory in the control group were higher than those in the exotropia group [(5.98±1.23)(6.80±1.53)，t=-2.81，P=0.01)]. In the reverse tapping test,the better the near stereopsis was,the higher the score was (B=0.78,95%CI=0.23-1.33,P=0.01),and the constant exotropia children performed better than the intermittent exotropia children(B=1.25,95%CI=0.16-2.24,P=0.03). Conclusion Visuospatial working memory is impaired in school-age children with exotropia,and the visuospatial working memory of exotropia children is affected by the near stereopsis and exotropia constancy.

Objective: To understand the characteristics of rapid naming in exotropia children, and to analyze the influence of clinical indicators related to exotropia on the rapid naming. Methods: A total of 45 exotropia children were recruited according to the diagnostic criteria of consensus of strabismus classification experts (2015) from the Zhongshan ophthalmic center as the case group, and 45 children of the same age, gender and parental educational status were recruited as the control group without any ocular diseases. All children were evaluated the ability of the rapid naming by classical rapid naming test. Results: The letter rapid naming time of children with exotropia was longer than that in control group [(26.87±10.18)(21.98±7.29)s], and the difference was statistically significant (t=2.73, P=0.01), however there was no significant correlation between strabismus degree, symptom duration, AC/A ratio, disease classification, simultaneous vision, the near stereopsis, the far stereopsis and the letter rapid naming in the clinical indicators of exotropia (r=-0.16, 0.23, 0.20, 0.06, 0.09, 0.05, 0.20, P>0.05). Conclusion Rapid naming might be impaired among children with exotropia, with no significant correlation between this defect and its clinical indicators.

OBJECTIVE: To assess the value of Ploton silver staining and phalloidin-iFlour 488 staining in observation of the morphology of osteocyte dendrites of mice at different developmental stages. METHODS: The humerus and femurs were harvested from mice at 0 (P0), 5 (P5), 15 (P15), 21 (P21), 28 (P28), and 35 days (P35) after birth to prepare cryo-sections and paraffin sections. HE staining of P35 mouse femur sections served as a reference for observing osteocytes in the trabecular bone and cortical bone. The humeral sections at different developmental stages were stained with Ploton silver staining to observe the morphology of osteocytes and canaliculi, and the canalicular lengths in the cortical and trabecular bones of the humerus of the mice in each developmental stage were recorded. The cryo-sections of the humerus from P10 and P15 mice were stained with phalloidin iFlour-488 to observe the morphology of osteocytes and measurement of the length of osteocyte dendrites in the cortical bone. RESULTS: In the trabecular bone of the humerus of P0-P15 mice, Ploton silver staining only visualized the outline of the osteocytes, and the morphology of the canaliculi was poorly defined. In P21 or older mice, Ploton silver staining revealed the morphology of the trabecular bone osteocytes and the canaliculi, which were neatly arranged and whose lengths increased significantly with age (P21 CONCLUSIONS Mouse osteocyte dendrites elongate progressively and their arrangement gradually becomes regular with age. Ploton silver staining can clearly visualize the morphology of the osteocytes and the canaliculi in adult mice but not in mice in early stages of development. Phalloidin iFlour-488 staining for labeling the cytoskeleton can be applied for mouse osteocytes at all developmental stages and allows morphological observation of mouse osteocytes in early developmental stages.
Animals ; Bone and Bones ; Dendrites ; Mice ; Osteocytes ; Phalloidine ; Silver Staining

Objective To explore the effect of tea polyphenols on the growth of human papillomavirus 16 (HPV16) subgenes-immortalized human cervical epithelial cells (H8 cells).Methods Cultured H8 cells were divided into 5 groups to be treated with 0 (control group),6.25,12.5,25 and 50 mg/L tea polyphenols respectively for 24,36,and 48 hours,and then cell counting kit-8 (CCK8)assay was performed to detect cell proliferation.After 24 hours of incubation,flow cytometry was conducted to detect cell apoptosis and cell cycle,and fluorescence microscopy to observe the morphology of apoptotic cells.Results After incubation with tea polyphenols at different concentrations for 24,36 and 48 hours,the proliferation of H8 cells was inhibited,and 12.5 mg/L tea polyphenols could inhibit the relative growth rate of H8 cells in a time-dependent manner.Flow cytometry showed that there was a significant difference in cell apoptosis rate among the 6.25-,12.5-,25-,50-mg/L tea polyphenols groups and the control group (52.62％ ± 0.62％,52.22％ ± 0.72％,42.52％ ± 0.90％,45.96％ ± 2.11％,29.96％ ± 0.70％ respectively,F =272.0,P ＜ 0.05).Moreover,all the tea polyphenol groups showed significantly increased cell apoptosis rate compared with the control group (all P ＜ 0.05).Fluorescence microscopy showed karyopyknosis,nuclear fragmentation and other typical apoptotic morphological changes in H8 cells in tea polyphenols groups.There were significant differences in the percentage of cells in G1,G2 phase and cell proliferation index among the 5 groups (all P ＜ 0.05).Compared with the control group,the 6.25-,12.5-,25-mg/L tea polyphenols groups showed significantly increased percentage of cells in G1 phase (55.96％ ± 0.72％,54.12％ ± 3.20％,65.30％ ± 1.51％ respectively,all P ＜ 0.05),but significantly decreased percentage of cells in G2 phase (3.17 ± 1.82％,4.94 ± 1.46％,4.65 ± 4.26％ respectively,all P ＜ 0.05) and lower cell proliferation index(0.44 ± 0.01,0.46 ± 0.02,0.36 ± 0.01 respectively,all P ＜ 0.05).Conclusion Tea polyphenols can inhibit the proliferation of H8 cells,induce cell apoptosis,and block cell cycle progression.

Objective To investigate the effect of ski gene in migration process of astrocytes in rats. Methods Astrocytes were obtained from rats' cerebral cortex and cultured in vitro. siRNA targeting ski gene and negative control sequences were prepared. The ski-siRNA group, siRNA negative control group and untreated group were set in this experiment. The specific siRNA targeting ski gene was transfected into astrocytes with Lipofectamine?RNAiMAX Reagent. Then the ski protein levels were determined with Western blotting. After transfec-tion, the changes in migration of astrocytes were measured with wound scratch assay and Transwell migration assay. Results Western blot-ting showed that the expression of ski protein was significantly lower in the ski-siRNA group than in the siRNA negative control group and untreated group (F=132.957, P<0.001). Transwell migration assay showed that the number of astrocytes crossing through chambers was less in the ski-siRNA group than in the siRNA negative control group and untreated group (F>47.197, P<0.05). Wound scratch assay showed that the wound healing rate was lower in the ski-siRNA group than in the control group one, two, three, four and five days after transfection (F>69.187, P<0.001). Conclusion Ski knocked down by siRNA could inhibit the migration ability of astrocytes. It is a reminding that ski may take part in the migration process of astrocytes, and moreover, ski may play an important role in the formation of glial scar.