Advanced Platelet-Rich Fibrin Extract Treatment Promotes the Proliferation and Differentiation of Human Adipose-Derived Mesenchymal Stem Cells through Activation of Tryptophan Metabolism.

BACKGROUND: Advanced platelet-rich fibrin extract (APRFE) contains a high concentration of various cytokines that are helpful for improving stem cells repair function. OBJECTIVE: However, the underlying mechanism of APRFE improving stem cell repairing is not clear. METHODS: We produced APRFE by centrifuging fresh peripheral blood samples and isolated and identified human adipose-derived mesenchymal stem cells (ADMSCs). The abundance of cytokines contained in APRFE was detected by the Enzyme-linked immunosorbent assay (ELISA). The ADMSCs treated with or without APRFE were collected for transcriptome sequencing. RESULTS: Based on the sequencing data, the expression profiles were contracted. The differentially expressed genes and lncRNA (DEGs and DElncRNAs) were obtained using for the differential expression analysis. The lncRNA-miRNA-mRNA network was constructed based on the miRNet database. The further enrichment analysis results showed that the biological functions were mainly related to proliferation, differentiation, and cell-cell function. To explore the role of APRFE, the protein-protein interaction network was constructed among the cytokines included in APRFE and DEGs. Furthermore, we constructed the global regulatory network based on the RNAInter and TRRUST database. The pathways in the global regulatory network were considered as the core pathways. We found that the DEGs in the core pathways were associated with stemness scores. CONCLUSION: In summary, we predicted that APRFE activated three pathways (tryptophan metabolism, mTOR signaling pathway, and adipocytokine signaling) to promote the proliferation and differentiation of ADMSCs. The finding may be helpful for guiding the application of ADMSCs in the clinic.

Analysis of online antenatal education class use via a mobile terminal app during the COVID-19 pandemic.

OBJECTIVE: To understand the use of online antenatal education classes accessed via the Mother and Child Health Handbook app during the COVID-19 pandemic in order to provide a basis and suggestions for optimizing Internet education during pregnancy under public health emergencies. METHODS: We compared and analyzed the use of online antenatal education classes via the Mother and Child Health Handbook app in Hangzhou in 2019 and 2020 (during the COVID-19 pandemic). RESULTS: Between January 1, 2019, and December 31, 2020, a total of 229,794 pregnant women created files and registered for the app, including 124,273 women in 2019 and 105,521 women in 2020. More pregnant women participated in online antenatal education learning (n = 36,379/34.5% vs. 29,226/23.5%, p = 0.000) in 2020 than in 2019. The proportion of pregnant women in the 18-34-year-old group who participated in online learning was higher than that in the advanced age group, and the difference was statistically significant (2019: 24.3% vs. 18.8%, p = 0.000) (2020: 35.7% vs. 27.4%, p = 0.000). More pregnant women accessed online antenatal education during early pregnancy (n = 13,463/37.0% vs. 9088/31.1%, p = 0.000) in 2020 than in 2019. Similar percentages of pregnant women participated in online antenatal education during mid-pregnancy (n = 15,426/52.8% vs. 19,269/53.0%, p = 0.639) in 2019 and 2020. Fewer pregnant women accessed online antenatal education during late pregnancy (n = 10,246/28.2% vs. 9476/32.4%, p = 0.000) in 2020 than in 2019. Fewer pregnant women choose to take 'Puerperal Health' courses in 2020 than in 2019 (early pregnancy: 36.20% vs. 42.79%, p = 0.000; mid-pregnancy: 41.65% vs. 48.19%, p = 0.000; late pregnancy: 55.31% vs. 58.41%, p = 0.000). Fewer pregnant women choose to take 'Psychological Adjustment' courses in 2020 than in 2019 (early pregnancy: 21.59% vs. 29.60%, p = 0.000; mid-pregnancy: 26.20% vs. 40.50%, p = 0.000; late pregnancy: 12.79% vs. 42.53%, p = 0.000). More pregnant women choose to study 'Nutrition and Exercise' in 2020 than in 2019 (early pregnancy: 44.48% vs. 25.95%, p = 0.000; mid-pregnancy: 47.77% vs. 40.75%, p = 0.000; late pregnancy: 55.94% vs. 42.99%, p = 0.000). "Pregnancy Care and Fetal Development" was the most selected course by pregnant women in early pregnancy (2019: 67.50%; 2020: 71.39%) and middle pregnancy (2019: 67.01%; 2020: 82.05%), and the proportion in 2020 was higher than it was in 2019. "Baby care" was the most selected course by pregnant women in late pregnancy, and the proportion in 2020 was higher than it was in 2019 (78.31% vs. 72.85%). CONCLUSION: During the COVID-19 pandemic, online antenatal education was well-used by pregnant women. More women participated in the online antenatal education modules during the COVID-19 pandemic than during 2019.The proportion of choosing different courses for pregnant women before and after the COVID-19 epidemic varied, and the learning course needs of pregnant women in different trimesters were different.

Utx Regulates the NF-κB Signaling Pathway of Natural Stem Cells to Modulate Macrophage Migration during Spinal Cord Injury.

Neural stem cells (NSCs) play vital roles in the homeostasis of neurological function. Ubiquitously transcribed tetratricopeptide repeat, X chromosome (UTX) is an important regulator of stem cell phenotypes. In our current study, we aimed to investigate whether the conditional knockout of UTX on neural stem cells alters macrophage assembly in response to spinal cord injury (SCI). Conditional knockout Utx of NSC (Utx-KO) mice was used to generate SCI models by the modified Allen method. We reported that neurological function and scar hyperplasia significantly improved in Utx-KO mice after SCI, accompanied by significantly reduced assembly of macrophages. With a 45-fold pathway array and Western blot, we found that Utx-KO could significantly inhibit NF-κB signaling activation and promote the synthesis and secretion of macrophage migration inhibitory factor (MIF) in NSCs. Administration of the selective NF-κB p65 activator betulinic acid and the selective MIF inhibitor ISO-1 confirmed that the activation of NF-κB p65 phosphorylation or inhibition of MIF could eliminate the benefits of Utx-KO in SCI, such as inhibition of macrophage aggregation and reduction in scar proliferation. This study confirmed that UTX in NSCs could alter macrophage migration and improve neurological function recovery after SCI in mice.

Overcoming Intrinsic H3K27me3 Imprinting Barriers Improves Post-implantation Development after Somatic Cell Nuclear Transfer.

Successful cloning by somatic cell nuclear transfer (SCNT) requires overcoming significant epigenetic barriers. Genomic imprinting is not generally regarded as such a barrier, although H3K27me3-dependent imprinting is differentially distributed in E6.5 epiblast and extraembryonic tissues. Here we report significant enhancement of SCNT efficiency by deriving somatic donor cells carrying simultaneous monoallelic deletion of four H3K27me3-imprinted genes from haploid mouse embryonic stem cells. Quadruple monoallelic deletion of Sfmbt2, Jade1, Gab1, and Smoc1 normalized H3K27me3-imprinted expression patterns and increased fibroblast cloning efficiency to 14% compared with a 0% birth rate from wild-type fibroblasts while preventing the placental and body overgrowth defects frequently observed in cloned animals. Sfmbt2 deletion was the most effective of the four individual gene deletions in improving SCNT. These results show that lack of H3K27me3 imprinting in somatic cells is an epigenetic barrier that impedes post-implantation development of SCNT embryos and can be overcome by monoallelic imprinting gene deletions in donor cells.

Role of Usual Healthcare Combined with Telemedicine in the Management of High-Risk Pregnancy in Hangzhou, China.

Background: Maternal health is an important part of basic public health services in China's medical reform. Effective management is an important guarantee of maternal health. Telemedicine has been widely used in maternal health management. Objective: This study explores the role of usual healthcare combined with telemedicine in the management of high-risk pregnancy. Methods: The study was a retrospective. Data were obtained from Hangzhou Maternity Hospital between October 2012 and September 2016, including 93465 pregnant women who were in usual high-risk pregnancy management (usual group) and 134884 pregnant women who were in telemedicine combined with usual high-risk pregnancy management (telemedicine group). The differences in high-risk scores and pregnancy outcomes between the usual and the telemedicine groups were compared. Results: The high-risk factors were analyzed, and the results showed that the first fixed high-risk factor was scar uterus and the first dynamic high-risk factor was hepatitis B. Comparing the data of two groups, the number of prenatal visits increased significantly in the telemedicine group (p value <0.05). Although the critical proportion of high-risk women was 2.13% in the usual group and 5.88% in the telemedicine group, respectively (p value <0.01), maternal mortality decreased in the telemedicine group (p value <0.05). Conclusion: The combination of telemedicine and usual healthcare can urge the pregnant women to carry out antenatal visits on time, which is one of the important factors to improve the outcome of high-risk pregnancy.

Silencing of lncRNA PKIA-AS1 Attenuates Spinal Nerve Ligation-Induced Neuropathic Pain Through Epigenetic Downregulation of CDK6 Expression.

Neuropathic pain (NP) is among the most intractable comorbidities of spinal cord injury. Dysregulation of non-coding RNAs has also been implicated in the development of neuropathic pain. Here, we identified a novel lncRNA, PKIA-AS1, by using lncRNA array analysis in spinal cord tissue of spinal nerve ligation (SNL) model rats, and investigated the role of PKIA-AS1 in SNL-mediated neuropathic pain. We observed that PKIA-AS1 was significantly upregulated in SNL model rats and that PKIA-AS1 knockdown attenuated neuropathic pain progression. Alternatively, overexpression of PKIA-AS1 was sufficient to induce neuropathic pain-like symptoms in uninjured rats. We also found that PKIA-AS1 mediated SNL-induced neuropathic pain by directly regulating the expression and function of CDK6, which is essential for the initiation and maintenance of neuroinflammation and neuropathic pain. Therefore, our study identifies PKIA-AS1 as a novel therapeutic target for neuroinflammation related neuropathic pain.

Arginine Vasopressin-Aquaporin-2 Pathway-Mediated Dehydration Effects of Electroacupuncture in Guinea Pig Model of AVP-Induced Eendolymphatic Hydrops.

OBJECTIVE: To investigate the effects of electroacupuncture (EA) on endolymphatic hydrops (EH) and the regulation of arginine vasopressin (AVP)-aquaporin-2 (AQP2) pathway in guinea pigs. METHODS: EH was induced in male guinea pigs by an intraperitoneal injection of AVP. For the treatment, EA was delivered to Baihui (GV 20) and Tinggong (SI 19) acupoints, once per day for 10 consecutive days. In histomorphological studies, cochlear hydrops degree was evaluated by hematoxylin-eosin (HE) staining, and then the ratio of scala media (SM) area to SM + scala vestibuli (SV) area (R value) was calculated. In mechanical studies, a comparison of plasma AVP (p-AVP) concentrations, cyclic adenosine monophosphate (cAMP) levels, vasopressin type 2 receptor (V2R) and AQP2 mRNA expressions in the cochlea were compared among groups. RESULTS: EA significantly reduced cochlear hydrops in guinea pigs (P=0.001). EA significantly attenuated the AVPinduced up-regulation of p-AVP concentrations (P=0.006), cochlear cAMP levels (P=0.003) and AQP2 mRNA expression (P=0.016), and up-regulated the expression of V2R mRNA (P=0.004) in the cochlea. CONCLUSIONS: The dehydrating effect of EA might be associated with its inhibition of AVP-AQP2 pathway activation.

METTL3-mediated m6A modification is required for cerebellar development.

N6-methyladenosine (m6A) RNA methylation is the most abundant modification on mRNAs and plays important roles in various biological processes. The formation of m6A is catalyzed by a methyltransferase complex including methyltransferase-like 3 (METTL3) as a key factor. However, the in vivo functions of METTL3 and m6A modification in mammalian development remain unclear. Here, we show that specific inactivation of Mettl3 in mouse nervous system causes severe developmental defects in the brain. Mettl3 conditional knockout (cKO) mice manifest cerebellar hypoplasia caused by drastically enhanced apoptosis of newborn cerebellar granule cells (CGCs) in the external granular layer (EGL). METTL3 depletion-induced loss of m6A modification causes extended RNA half-lives and aberrant splicing events, consequently leading to dysregulation of transcriptome-wide gene expression and premature CGC death. Our findings reveal a critical role of METTL3-mediated m6A in regulating the development of mammalian cerebellum.

Sacroiliac joint tuberculosis: surgical management by posterior open-window focal debridement and joint fusion.

BACKGROUND: Sacroiliac joint tuberculosis(SJT) is relatively uncommon, but it may cause severe sacroiliac joint destruction and functional disorder. Few studies in the literature have been presented on SJT, reports of surgical treatment for SJT are even fewer. In this study, we retrospectively reviewed surgical management of patients with severe SJT of 3 different types and proposed to reveal the clinical manifestations and features and aim to determine the efficiency and security of such surgical treatment. METHODS: We reviewed 17 patients with severe SJT of 3 different types who underwent posterior open-window focal debridement and bone graft for joint fusion. Among them,five patients with anterior sacral abscess had anterior abscess curettage before debridement. Two patients with lumbar vertebral tuberculosis received one-stage posterior tuberculous debridement, interbody fusion and instrumentation. Follow-up was performed 36 months (26 to 45 months) using the following parameters: erythrocyte sedimentation rate(ESR), status of joint bony fusion on CT scan, visual analogue scale (VAS) and the Oswestry Disability Index (ODI). RESULTS: Buttock pain and low back pain were progressively relieved with time. 6 months later, pain was not obvious, and ESR resumed to normal levels within 3 months. Solid fusion of the sacroiliac joint occurred within 12 months in all cases. No complications or recurrence occurred. At final follow-up, all patients had no pain or only minimal discomfort over the affected joint and almost complete functional recovery. CONCLUSIONS: Posterior open-window focal debridement and joint fusion is an efficient and secure surgical method to treat severe SJT. If there is an abscess in the front of the sacroiliac joint, anterior abscess curettage should be performed as a supplement.

Mettl3-mediated m6A regulates spermatogonial differentiation and meiosis initiation.

METTL3 catalyzes the formation of N6-methyl-adenosine (m6A) which has important roles in regulating various biological processes. However, the in vivo function of Mettl3 remains largely unknown in mammals. Here we generated germ cell-specific Mettl3 knockout mice and demonstrated that Mettl3 was essential for male fertility and spermatogenesis. The ablation of Mettl3 in germ cells severely inhibited spermatogonial differentiation and blocked the initiation of meiosis. Transcriptome and m6A profiling analysis revealed that genes functioning in spermatogenesis had altered profiles of expression and alternative splicing. Our findings provide novel insights into the function and regulatory mechanisms of Mettl3-mediated m6A modification in spermatogenesis and reproduction in mammals.

Crystal structure of (5'S,8'S)-3-(2,5-di-methyl-phen-yl)-8-meth-oxy-3-nitro-1-aza-spiro-[4.5]decane-2,4-dione.

The title compound, C18H22N2O5, was synthesized by nitrification of its enol precursor. The pyrrolidine ring plane adopts a twisted conformation about the C-C bond linking the spiro centre and the C=O group remote from the N atom. It makes dihedral angles of 71.69 (9) and 88.92 (9)°, respectively, with the benzene ring plane and the plane defined by the four C atoms that form the seat of the of the cyclo-hexane chair. At the spiro centre, the NH group is axial and the C=O group is equatorial with respect to the cyclo-hexane ring. In the crystal, inversion dimers linked by pairs of N-H⋯O hydrogen bonds generate R 2 (2)(8) loops. The dimers are linked by C-H⋯O inter-actions, generating a three-dimensional network.

Simultaneous quantitation of folic acid and 5-methyltetrahydrofolic acid in human plasma by HPLC-MS/MS and its application to a pharmacokinetic study.

A sensitive method based on high-performance liquid chromatography-tandem mass spectrometry (LC-MS/MS) has been developed for the simultaneous determination of folic acid (FA) and its active metabolite, 5-methyltetrahydrofolic acid (5-M-THF), in human plasma. The analytes were extracted from plasma with methanol solution containing 10 mg/mL of 2-mercaptoethanol and 0.025% (v/v) ammonium hydroxide. FA and 5-M-THF were more stable after the addition of 2-mercaptoethanol and ammonium hydroxide in the sample preparation procedures of this study than they were in the previously published methods. Chromatographic separation was performed on a Hedera ODS-2 column using a gradient elution system of acetonitrile and 1 mM ammonium acetate buffer solution containing 0.6% formic acid as mobile phase. LC-MS/MS was carried out with an ESI ion-source and operated in the multiple reaction monitoring (MRM) mode. The assay was linear over the concentration ranges of 0.249-19.9 ng/mL for FA, and 5.05-50.5 ng/mL for 5-M-THF. The developed LC-MS/MS method offers increased sensitivity for quantification of FA and 5-M-THF in human plasma and was applicable to a pharmacokinetic study of FA and 5-M-THF.

[LC-MS/MS assay of methylphenidate: stability and pharmacokinetics in human].

The study aims to solve the instability problem of methylphenidate (MPH) in plasma, and establish a LC-MS/MS method for simultaneous determining of MPH in human plasma. The stabilities of MPH in different media were studied, and the degradation characteristics of MPH in these media were also investigated by HPLC and LC-MS/MS. To a 200 microL aliquot of freshly collected plasma sample, 10 microL 2% formic acid was added immediately to prevent the hydrolysis of MPH in human plasma samples. Chromatographic separation was performed on a Sapphire C18 column using the mobile phase of methanol - 5 mmol.L-1 ammonium acetate buffer solution containing 0.1% formic acid (46 : 54). MPH was quantified by tandem mass spectrometry operating in positive electrospray ionization mode with multiple reaction monitoring. The detection used the transitions of protonated molecules at m/z 234.2-->84.1 for MPH and m/z 260.3-->183.1 for propranolol (IS), separately. The intra- and inter-assay precisions were all below 5.0%. The accuracies were all in standard ranges. The linear calibration curve was obtained in the concentration range of 0.035-40 ng.mL-1. The methods fulfilled the demand. The method was used to determine the concentration of MPH in human plasma after a single dose of 36 mg MPH tablet to 6 healthy Chinese volunteers. The method is suitable for the precisely determination of MPH and for pharmacokinetic study of MPH in human plasma.

[Drug resistance of methicillin resistant Staphylococcus strains in burn ward and relative analysis].

OBJECTIVE: To study the distribution and drug resistance of methicillin resistant Staphylococcus strains in various specimens of inpatients in burn wards, and to provide reference for clinical treatment. METHODS: Bacteria were isolated from specimens of wound exudate, blood, sputum, and bronchoalveolar lavage fluid etc., which were collected from patients hospitalized in our burn wards from January 2008 to December 2010. The bacteria were routinely cultured and identified. Drug resistance of the Staphylococci to 15 antibiotics commonly used in clinic was identified by K-B disk diffusion method. Data were processed with statistical software WHONET 5.5. The homology of 40 strains of methicillin resistant Staphylococcus aureus (MRSA) was analyzed by pulsed-field gel electrophoresis (PFGE). RESULTS: Altogether 386 strains of Staphylococcus were isolated, including 196 strains of Staphylococcus aureus and 190 strains of coagulase negative Staphylococcus. The mean annual isolation rates of MRSA and methicillin resistant coagulase negative Staphylococcus (MRCoNS) were respectively 73.00% (143/196) and 74.20% (141/190). The resistance rates of MRSA and MRCoNS to ß-lactams drugs, such as penicillin, oxacillin, cefazolin, and cefuroxime were 100.00% in every year. No Staphylococcus strains resistant to vancomycin, teicoplanin, or linezolid were found. Three different PFGE patterns A, B, and C were identified among 40 MRSA strains, including 33 strains of type A (30 strains in sub-type A1 and 3 strains in sub-type A2), 6 strains of type B (respectively 3 strains in sub-types B1 and B2), and 1 strain of type C. CONCLUSIONS: The isolation rates of MRSA and MRCoNS were high in our burn wards from January 2008 to December 2010. All of them showed strong drug resistance property, and they were multidrug resistant. The most prevalent strain was PFGE type A.

[Detection of integron, homology and genotype of carbapenemases in multi-drug resistant Acinetobacter Baumannii isolated from burn wound].

OBJECTIVE: To investigate the relationship among antibiotic resistance, integron, homology of multi-drug resistant Acinetobacter Baumannii isolated from burn ward. METHODS: Thirty-one strains of multi-drug resistant Acinetobacter baumannii were isolated from samples of burn wound exudate in hospitalized patients of Gansu Province People's Hospital. The minimum inhibitory concentrations (MIC) of these strains against 11 antibiotics was examined by agar dilution method. Homology of these strains was analyzed by pulse-field gel electrophoresis (PFGE). Class 1, 2 and 3 integrase, integron genes and genotype of carbapenemases were amplified by PCR and verified by DNA sequencing. RESULTS: Acinetobacter baumannii were highly resistant to all antibiotics except imipenem, meropenem, cefoperazone-sulb-actam, piperacillin-tazobactam (antibiotic resistance rate was 45.2%, 48.4%, 48.4%, 41.0%, respectively). All strains were classified into 3 types of clones (A, B, C clone included 18, 7, 6 strains respectively) based on PFGE pattern. Integrons of 20 strains of Acinetobacter Baumannii harbored aadA1, aadA5, aacA4, aac3, catB8, aacC1, aac (6')-Ib, drfA17 and drf8 gene. CONCLUSION: Multi-drug resistance Acinetobacter baumannii (major in clone A) spread widely in burn ward of Gansu Province People's Hospital. Integrons of Acinetobacter baumannii mediated drug resistance against aminoglycoside antibiotics, chloramp-phenicol. All carbapenem-resistant Acinetobacter Baumannii can produce OXA-23 carbapenemase.

[Synthesis and antithrombotic activity of acetylsalicyl ferulic acid-coupling furoxans and nitrates].

AIM: To synthesize and study the antithrombotic activity of NO-donating aspirin derivatives. METHODS: Furoxans and nitrates were incorporated to aspirin via antioxidant ferulic acid as a linker, and the target compounds were screened for in vitro and in vivo inhibitory activities of platelet aggregation, and for inhibitory effect on A-V hypass thromhosis in rats. RESULTS: Fourteen novel compounds I(1-14), were synthesized and their structures were confirmed Iy MS, IR, 1H NMR and elemental analysis. Biological screening results demonstrated that some tested compounds exhibited potential antithrombotic activ it. CONCLUSION: Acetylsalicyl ferulic acid-coupling furoxans and nitrates might he used as a lead for further study.