RESUMO
Clinical application of doxorubicin (DOX) is heavily hindered by DOX cardiotoxicity. Several theories were postulated for DOX cardiotoxicity including DNA damage and DNA damage response (DDR), although the mechanism(s) involved remains to be elucidated. This study evaluated the potential role of TBC domain family member 15 (TBC1D15) in DOX cardiotoxicity. Tamoxifen-induced cardiac-specific Tbc1d15 knockout (Tbc1d15CKO) or Tbc1d15 knockin (Tbc1d15CKI) male mice were challenged with a single dose of DOX prior to cardiac assessment 1 week or 4 weeks following DOX challenge. Adenoviruses encoding TBC1D15 or containing shRNA targeting Tbc1d15 were used for Tbc1d15 overexpression or knockdown in isolated primary mouse cardiomyocytes. Our results revealed that DOX evoked upregulation of TBC1D15 with compromised myocardial function and overt mortality, the effects of which were ameliorated and accentuated by Tbc1d15 deletion and Tbc1d15 overexpression, respectively. DOX overtly evoked apoptotic cell death, the effect of which was alleviated and exacerbated by Tbc1d15 knockout and overexpression, respectively. Meanwhile, DOX provoked mitochondrial membrane potential collapse, oxidative stress and DNA damage, the effects of which were mitigated and exacerbated by Tbc1d15 knockdown and overexpression, respectively. Further scrutiny revealed that TBC1D15 fostered cytosolic accumulation of the cardinal DDR element DNA-dependent protein kinase catalytic subunit (DNA-PKcs). Liquid chromatography-tandem mass spectrometry and co-immunoprecipitation denoted an interaction between TBC1D15 and DNA-PKcs at the segment 594-624 of TBC1D15. Moreover, overexpression of TBC1D15 mutant (∆594-624, deletion of segment 594-624) failed to elicit accentuation of DOX-induced cytosolic retention of DNA-PKcs, DNA damage and cardiomyocyte apoptosis by TBC1D15 wild type. However, Tbc1d15 deletion ameliorated DOX-induced cardiomyocyte contractile anomalies, apoptosis, mitochondrial anomalies, DNA damage and cytosolic DNA-PKcs accumulation, which were canceled off by DNA-PKcs inhibition or ATM activation. Taken together, our findings denoted a pivotal role for TBC1D15 in DOX-induced DNA damage, mitochondrial injury, and apoptosis possibly through binding with DNA-PKcs and thus gate-keeping its cytosolic retention, a route to accentuation of cardiac contractile dysfunction in DOX-induced cardiotoxicity.
RESUMO
ObjectiveTo explore the differences existing in the auditory mismatch negativity （MMN） amplitude and latency between children with attention deficit hyperactivity disorder （ADHD） and normal children， and to probe into the significance of MMN latency and amplitude for assessing the auditory perception and attention level in ADHD children and normal children. MethodsOn December 1， 2022， a systematic search was performed in PubMed， Embase， Cochrane Library， China National knowledge Infrastructure （CNKI）， Wanfang Data Knowledge Service Platform and VIP databases to identify all well qualified literature focusing on MMN of ADHD children， then the valid data relevant to MMN amplitude and latency were extracted. The Newcastle-Ottawa Scale （NOS） was used to assess the quality of the included studies， and Stata 20.0 was employed for Meta-analysis. ResultsA total of 9 qualified studies comparing ADHD children （n=170） against healthy controls （n=159） were finally included. Among the included literature， there were 18 matched pairs of MMN amplitude data and 10 matched pairs of MMN latency data at different recording sites. Meta-analysis denoted that ADHD group resulted in potentials of slightly lower MMN amplitude （WMD=-0.334， 95% CI： -1.426~0.758， P=0.549） and notably longer MMN latency （WMD=14.768， 95% CI： 4.660~24.876， P=0.004） compared to control group， and the Bgger's funnel plot did not reveal any publication bias. ConclusionCompared with healthy controls， ADHD children have longer MMN latency， suggesting that the auditory perception and attention level of ADHD children may be reduced.
RESUMO
Objective:To establish Crx-iCreERT2 fluorescent reporter human embryonic stem cell lines using CRISPR/Cas9 technology and 3D retinal organoid culture.Methods:The target site sequence of H9 cell line was verified by polymerase chain reaction (PCR). SgRNAs were designed by CRISPR/Cas9 technique and their activity was detected.The most optimal sgRNA was selected according to the factors such as activity and specificity.After identification of the target vectors by restriction enzyme and sequencing, the target vectors were transferred to the H9 cell line by electroporation.P2A-tdTomato-P2A-iCreERT2 was inserted between Exon4 and 3’-untranslated region of hES-ZLM-001 gene.Knockin positive clones were obtained after drug treatment, enrichment of positive clones.Primers were designed to perform PCR on the target region, and homozygous de-resistant knockin positive cell clones were selected according to the sequencing results and peaks.The 1-A07 cell line was cultured, and then flow cytometry for the proportion of OCT4 positive cells, immunofluorescence for three stem cell molecular markers including SOX2, NANOG, SSEA4, karyotype analysis were carried out to confirm whether the 1-A07 cell line could be used for further experiments.Retinal organoids were obtained by three-dimensional (3D) culture technology and the expression of molecular markers was detected by immunofluorescence at different developmental stages of retinal organoids. Results:The target site sequence of H9 cell line was consistent with that given by Genebank and Ensembl.Sixteen sgRNAs were designed according to the target site sequence of H9 cell line, and finally sgRNA8 and sgRNA12 were selected.The sgRNAs and recombinant plasmids were transfected into the H9 cell line by electroporation, and four homozygous de-resistant knockin positive cell clones were obtained by PCR.Crx-iCreERT2 fluorescent reporter human embryonic stem cell lines were successfully obtained.In 1-A07 cell line, the proportion of OCT4 positive cells was about 98.7% by flow cytometry, and the expression of three stem cell markers was positive by immunofluorescence, and the karyotype was normal 46, XX.The results showed that the 1-A07 cell line could be used for further experiments.The Crx-iCreERT2 fluorescent reporter human embryonic stem cell lines were differentiated into tdTomato positive retinal organoids by 3D culture technology.BRN3A positive ganglion cells, CALBINDIN positive horizontal cells and CHAT positive amacrine cells appeared on day 30 of differentiation.RECOVERIN positive photoreceptors arose on day 45 of differentiation.PKCα positive bipolar cells presented on day 90 of differentiation.Ganglion cells were shown in the deep layer of retinal organoids, and horizontal cells, amacrine cells and bipolar cells in the middle layer, and photoreceptors in the top layer.Conclusions:Crx-iCreERT2 fluorescent reporter human embryonic stem cell lines are successfully established and can be differentiated into retinal organoids that express tdTomato red fluorescence through 3D culture technology.Those retinal organoids contain the same types of neurons as normal human retinas, and follow a certain temporal and spatial developmental sequence similar to the developmental rules of normal human retinas.Crx-iCreERT2 fluorescent reporter human embryonic stem cell line is a powerful tool for researching retinal development and diseases and can be applied in treatments for blindness.
RESUMO
ObjectivesThe main aim of our study is to observe clinical characteristics and effects of antihypertensive drugs in different hospitalized populations, and to further provide evidence to explore causes and specific clinical markers of the aggravation of COVID-19 in patients with underlying hypertension. DesignThis was a retrospective cohort study focusing on the clinical data of COVID-19 inpatients admitted at the early stage of pandemic. SettingA single center study conducted in Tongji Hospital, Tongji Medical College of Huazhong university of Science and Technology (Wuhan, China). ParticipantsAll 1031 inpatients diagnosed with COVID-19 according to Prevention and control Scheme for Novel Coronavirus Pneumonia published by National Health Commission of the Peoples Republic of China and WHO interim guidance in Tongji hospital (Wuhan, China), from January 27, 2020, to March 8, 2020 with the cutoff date being March 30, 2020, were included in this study. Main outcome measuresDemographic data, medical history, clinical symptoms and signs, laboratory findings, chest computed tomography (CT), treatment, and clinical outcomes were extracted from electronic medical records. Results1031 COVID-19 inpatients were included in this study, of whom 866 were discharged and 165 were deceased in hospital. 73% of 165 deceased patients had chronic comorbidities. Patients with underlying diseases showed CFR 2.8 times as that of patients without. Senility and males were observed to be main risk factors for increased in-hospital case-fatality rate, with the odds ratio in multivariable regression being 2.94 (95%CI: 2.00 to 4.33; P <0.001) and 2.47 (95%CI: 1.65 to 3.70; P <0.001), respectively. The odds ratio of cases with composite endpoints for patients with simple hypertension was 1.53 (95%CI: 1.07 to 2.17; P=0.019). Senile patients with hypertension were proved to be at high risk early in the disease, which might be associated with the level of CRP, LDH, and eGFR. The odds ratio of case-fatality rate for patients with hypertension taking CCB group was 0.67 (95%CI: 0.37 to 1.20; P = 0.176). Among 271 severe cases without IKF, the odds ratio of case-fatality rate was 0.42 (95CI%: 0.18 to 0.99; P = 0.046) for patients in the CCB group after adjustment of age, sex, and underlying diseases. ConclusionsHypertension is not just a chronic underlying comorbidity, but also a risk factor affecting the severity of COVID-19 and does play a critical role in worsening patients clinical outcomes. Therefore, hypertension management in patients with COVID-19 should be regarded as a major challenge in the diagnostic and therapeutic strategies. Trial registrationN.A.
RESUMO
The current status and research progress of sanitation of public articles and appliances were described on the basis of the recent literature reviews.The researches on methods for disinfection of public articles and appliances showed that the most efficient method for disinfection of haircut appliance was ultraviolet radiation disinfection.The disinfetion efficiencies of different methods for disinfection of tea sets,towels,etc,the recent new reports on the selection of sanitary parameters and sampling methods of sanitary standard for public articles were introduced also.
RESUMO
Objective To understand the efficiency of defluoridation facilities used in drinking water defluoridation in the rural high fluoride areas in Tianjin and the sanitary characteristics of treated drinking water. Methods The raw water samples and treated water samples were collected from 4 water stations where the electrodialysis technics was applied and 6 water stations where the reverse osmosis technics was applied and the facilities had been operated for 2-3 years in a county, as well as 3 water stations applied activated aluminium oxide adsorption technics and 2 water stations applied nanofiltration membrane technics in the other counties in Tianjin in 2002-2004. The pH value,turbidity, total dissolved solids, total hardness, fluoride, sulfate, aluminium, in all water samples were determined and evaluated according to the Sanitary Standard for Drinking Water(2001). Results The treated water samples from electrodialysis and reverse osmosis defluoridation facilities showed lower fluoride level(below 1.0 mg/L with fluoride removal rates of 90.01% and 75.32%),and lower total dissolved solids and total hardness levels with removal rates of 75.83%-93.48%. The treated water samples from nanofiltration membrane defluoridation facilities also showed lower fluoride level(below 1.0 mg/L with fluoride removal rates of 75.16%,and lower total dissolved solids and total hardness levels with removal rates of 64.90% and 77.22% respectively. Conclusion The investigated electrodialysis, reverse osmosis and nanofiltration membrane defluoridation facilities show a reliable efficiency and are suitable for application of removal of fluoride in drinking water with different levels of total dissolved solids in the rural high fluoride areas. The effects of defluoridated water produced by reverse osmosis defluoridation facilities on human health need to be studied further more.
