ObjectiveTo observe the effects of Dihuang Yinzi （DY） on motor dysfunction in rats with advanced Parkinson's disease （PD） and to investigate the mechanisms by which DY improves advanced PD symptoms through the "gut microbiota-short-chain fatty acids （SCFAs）-inflammation-neuroprotection pathway". MethodsAn advanced PD rat model was induced by rotenone. Rats were divided into a normal group， model group， positive drug group （levodopa， 50 mg·kg^-1）， and DY low-， medium-， and high-dose groups （5.2， 10.4， 20.8 g·kg^-1）. After 7 days of administration， motor function was evaluated using the open-field， pole-climbing， and inclined plate tests. Hematoxylin-eosin （HE） staining was used to observe pathological changes in the substantia nigra and colon， and immunohistochemistry was performed to detect α-Synuclein （α-Syn） and tyrosine hydroxylase （TH） expression in the substantia nigra. Enzyme-linked immunosorbent assay （ELISA） was used to measure levels of dopamine （DA）， 5-hydroxytryptamine （5-HT）， 3，4-dihydroxyphenylacetic acid （DOPAC）， Levodopa， homovanillic acid （HVA）， tumor necrosis factor-α （TNF-α）， interleukin-6 （IL-6）， and interleukin-1β （IL-1β）. Western blot analysis was used to detect the expression of zonula occludens-1 （ZO-1） and occludin. Gut microbiota diversity was analyzed by 16S rRNA sequencing， and gas chromatography （GC） was used to determine the content of SCFAs in colonic contents. ResultsCompared with the normal group， the model group showed significantly decreased movement speed and distance in the open-field test， prolonged pole-climbing time， and reduced retention angle on the inclined plate （P<0.01）， accompanied by increased α-Syn expression （P<0.01） and decreased TH expression （P<0.01） in the brain. Compared with the model group， all DY dose groups improved motor dysfunction in advanced PD rats to varying degrees （P<0.05， P<0.01） and alleviated pathological damage in the brain and colon. High-dose DY significantly reduced α-Syn aggregation in the substantia nigra （P<0.01） and increased TH expression （P<0.01）. ELISA and Western blot results showed that， compared with the normal group， the model group exhibited decreased levels of DA， 5-HT， DOPAC， Levodopa， and HVA in the striatum （P<0.01）， increased levels of TNF-α， IL-6， and IL-1β in the colon and striatum （P<0.01）， and significantly reduced expression of ZO-1 （P<0.05） and occludin in the colon （P<0.01）. Compared with the model group， all DY dose groups increased the levels of DA， 5-HT， DOPAC， Levodopa， and HVA in the striatum to varying degrees （P<0.05， P<0.01）. In the high-dose DY group， the levels of TNF-α， IL-6， and IL-1β in the colon and striatum were reduced （P<0.01）， while the expression of ZO-1 （P<0.05） and occludin in the intestine was increased. The 16S rRNA sequencing results indicated that the relative abundances of Actinobacteriota， Enterobacteriaceae， and Erysipelotrichaceae were increased in the model group， whereas the relative abundances of Bacteroidota， class Clostridia， Lachnospiraceae， and Akkermansia muciniphila were decreased. These changes were effectively reversed after high-dose DY intervention. GC analysis showed that the content of SCFAs in the colonic contents of rats in the model group was decreased （P<0.05， P<0.01）， while after high-dose DY intervention， the levels of acetate， propionate， isobutyrate， and butyrate were significantly increased （P<0.05， P<0.01）. ConclusionDY may exert therapeutic effects in advanced PD by regulating the gut microbiota-SCFAs-inflammation pathway.

ObjectiveTo observe the intervention effect of Fufang Kangjiaolv capsules on anxiety-like behaviors in the rat model of chronic restraint stress （CRS） and explore the mechanism underlying the anti-anxiety effect via the microbiota-gut-brain axis. MethodsRats were assigned into blank， model， positive drug （diazepam， 1 mg·kg^-1）， and low-， medium-， and high-dose （0.75， 1.5， 3 g·kg^-1， respectively） Fufang Kangjiaolv capsules groups. After 14 days of administration， the elevated plus maze test， open field test， light and dark box test， and marble burying test were performed. Hematoxylin-eosin staining was employed to observe the pathological changes in the hippocampus and colon of rats， and Nissl staining was conducted to observe the damage of hippocampal neurons. The gut microbiota was analyzed by 16S rRNA gene sequencing. Real-time fluorescence quantitative polymerase chain reaction （Real-time PCR） was employed to determine the mRNA levels of zonula occludens-1 （ZO-1） and occludin in the colon of rats. The levels of tumor necrosis factor-α （TNF-α）， interleukin-6 （IL-6）， and interleukin-1β （IL-1β） in the colon， serum， and hippocampus were determined by enzyme-linked immunosorbent assay. Western blot was employed to determine the protein levels of ZO-1， occludin， nuclear factor-κB p65 （NF-κB p65） in the colon tissue and NF-κB p65 and brain-derived neurotrophic factor （BDNF） in the hippocampal tissue. ResultsCompared with the blank group， the model group showed reductions in the time and frequency ratio of rats entering the elevated plus maze， the time and frequency of rats entering the central area of the open field， the time of entering the open box， the times of passing through the light and dark box， and the number of unburied beads （P<0.05， P<0.01）. Compared with the model group， Fufang Kangjiaolv capsules ameliorated the anxiety of the model rats to varying degrees， and the high-dose group had the best effect， with increases in the proportions of time and frequency of rats entering the open arm in the elevated plus maze （P<0.05）， the number of rats entering the central area in the open field （P<0.05）， the time of entering the open box， the times of passing through the light and dark boxes， and the number of unburied beads （P<0.01）. Moreover， the high-dose group showed alleviated pathological damage of hippocampal neurons and colon. The results of 16S rRNA gene sequencing showed that the model group had increased relative abundance of Firmicutes， Deferribacterota， Romboutsia， and Phascolarctobacterium， while it had decreased relative abundance of Bavcteroidota and Lactobacillus. The drug administration groups showed increased relative abundance of Bavcteroidota， Bacteroides， norank f norank o Clostridia UCG-014， and Blautia and decreased relative abundance of Firmicutes and Deferribacterota. Compared with the blank group， the model group showed down-regulated protein and mRNA levels of ZO-1 and occludin in the colon （P<0.01）， elevated levels of TNF-α， IL-6， and IL-β in the colon， serum， and hippocampus （P<0.01）， up-regulated protein level of NF-κB p65 in the colon and hippocampus （P<0.01）， and down-regulated protein level of BDNF in the hippocampus （P<0.05）. Compared with the model group， high-dose Fufang Kangjiaolv capsules up-regulated the mRNA levels of ZO-1 and occludin in the colon （P<0.01）， lowered the levels of TNF-α， IL-6， and IL-β in the colon， serum， and hippocampus （P<0.01）， up-regulated the protein levels of ZO-1 （P<0.01） and occludin （P<0.05） in the colon， down-regulated the protein level of NF-κB p65 in the colon and hippocampus （P<0.05）， and up-regulated the protein level of BDNF in the hippocampus. ConclusionFufang Kangjiaolv capsules can reduce the anxiety-like behaviors in the rat model of CRS by regulating the gut microbiota disturbance， up-regulating the expression of tight junction proteins in the colon， repairing intestinal mucosal mechanical barrier， and down-regulating NF-κB/BDNF signaling pathway， thereby reducing peripheral and central inflammation. This study proves the hypothesis that Fufang Kangjiaolv capsules play an anti-anxiety role via the microbiota-gut-brain axis， providing a new idea for further research.

Objective:To explore the characteristics of SMN1 gene variants and carry out functional verification for two children with Spinal muscular atrophy (SMA). Methods:Two male children with complicated SMA diagnosed at the Children′s Hospital Affiliated to Capital Institute of Pediatrics respectively in July 2021 and April 2022 due to delayed or retrograde motor development were selected as the study subjects. Clinical data of the children were collected. Primary culture of skin fibroblasts was carried out, and peripheral blood samples were collected from both children and their parents. Multiplex ligation-dependent probe amplification, combined long-range PCR and nested PCR, and Sanger sequencing were carried out to detect the copy number and variants of the SMN1 gene. Absolute quantitative real-time PCR, Western blotting and immunofluorescence were used to determine the transcriptional level of the SMN gene, expression of the SMN protein, and the number of functional SMN protein complexes (gems body), respectively. This study was approved by Medical Ethics Committee of the Children′s Hospital Affiliated to Capital Institute of Pediatrics (Ethics No. SHERLLM2021009). Results:Child 1, a 1-year-old boy, was clinically diagnosed with type 1 SMA. Child 2, a 2-and-a-half-year-old boy, was clinically diagnosed with type 3 SMA. Both children were found to harbor a paternally derived SMN1 deletion and a maternally derived SMN1 gene variant, namely c. 824G>T (p.Gly275Val) and c. 884A>T (p.*295Leu). Compared with the normal controls and carriers, the levels of full-length SMN1 transcripts in their peripheral blood and skin fibroblast cell lines were significantly decreased ( P<0.05), and the levels of SMN protein normalized to that of β-actin, and the numbers of gems bodies in the primary fibroblast cells were also significantly lower ( P<0.05). Based on the guidelines from the American College of Medical Genetics and Genomics, both variants were classified as likely pathogenic (PS3+ PM3+ PM5+ PP3; PS3+ PM3+ PM4+ PP3). Following the diagnosis, both children had received nusinersen treatment. Although their motor function was improved, child 1 still died at the age of 2 due to severe pulmonary infection. The walking ability of child 2 was significantly improved, and his prognosis appeared to be good. Conclusion:Two cases of clinically complicated SMA have been confirmed by genetic testing and experimental studies, which has provided a reference for their accurate treatment.

Objective To investigate the clinical and cytogenetic characteristics of acute myeloid leukemia(AML)patients with NRAS mutations.Methods Newly diagnosed AML patients in our hospital from January 2020 to August 2022 were selected,and their clinical data were retrospectively analyzed.According to NRAS mutations,the patients were divided into NRAS mutation group and NRAS wild group.The clinical characteristics and cytogenetic differences were compared between the two groups.Results A total of 162 newly diagnosed AML patients were included in this study.There were 28 in NRAS mutation group and 134 in NRAS wild group.The peripheral white blood cell count of NRAS mutation group was significantly higher than that of NRAS wild type group(53.10×109/L vs 24.78×109/L,P<0.05).There were no significant differences in the hemoglobin level,platelet count or bone marrow blast cell count between the two groups(P>0.05).The coexisting gene mutation occurred in 25 patients(89.3%,25/28)in NRAS mutation group.The most common coexisting gene mutation was KRAS,with a mutation rate of 28.6%.Compared with NRAS wild group,NRAS mutation group was more likely to obtain KRAS mutations(P<0.05).There was no significant difference in other coexisting mutated genes between the two groups(P>0.05).The proportion of poor prognosis karyotype in the NRAS mutation group was 23.1%,which was significantly higher than that in NRAS wild group(P<0.05).The proportions of favorable and intermediate prognosis karyotypes in NRAS mutation group were 7.7%and 69.2%,respectively,which were not significantly different from those in NRAS wild group(P>0.05).Conclusion The incidence of NRAS mutation is 17.3%in AML patients in this study.Patients with NRAS mutation are more likely to have KRAS mutation and have a higher proportion of poor prognosis karyotype.

Objective:To explore the distribution of the copy number of survival motor neuron gene 2 ( SMN2) and the transcript level of the full-length SMN2 ( fl-SMN2) transcript level in patients with type 1-3 spinal muscular atrophy (SMA), and to evaluate their influences on disease severity, progression, and prognosis. Methods:It was a retrospective study involving 78 therapy-naive SMA patients with SMN1 gene homozygous deletion who were diagnosed and treated in the Capital Institute of Pediatrics from January 2019 to December 2021.Cross-sectional clinical data, including age at onset, motor milestones, and complications were recorded.They were followed up for monitoring motor function degeneration and survival.The copy number of SMN2 and the transcript level of fl-SMN2 were detected.Differences between groups were compared by the Student′s t-test or One- Way ANOVA or Chi- square test.Kaplan-Meier analysis was used for survival analysis, and Kendall′ s tau- c was performed to assess the correlation of these two biomarkers with SMA phenotypes, age at onset, motor milestones, and survival. Results:Of the 78 SMA patients, there were 17 cases (21.8%) of type 1, 34 cases(43.6%) of type 2, and 27 cases(34.6%) of type 3.Seven cases(41.2%) type 1 SMA patients died, with a median survival time of 11 months, and no deaths were observed in type 2 and type 3 SMA patients.There was a significant difference in the median age at onset among SMA patients with 2, 3, and 4 copies of SMN2 (1.8, 12.0, and 24.0 months, respectively; F=4.943, P=0.01). The mean transcript level of fl-SMN2 in type 1, 2 and 3 SMA patients were 196.25±68.79, 331.21±108.79 and 455.69±122.27, respectively ( F=37.154, P<0.001). The survival rate of SMA with 2 SMN2 copies at 1, 2, and 5 years were 50.5%, 0, and 0, respectively, and their median survival age was 7 months.The survival rate of SMA with 3 and 4 SMN2 copies at 5 years were 97.4% and 100.0%, respectively.Moreover, a negative correlation was observed between the transcript level of fl-SMN2 and phenotype severity ( Kendall′ s tau- c=-0.444, P<0.001), and the transcript level of fl-SMN2 of the survival group was much higher than that of the death group (342.93±125.74 vs.212.14±92.31). More copies of SMN2 and higher transcript level of fl- SMN2 indicated more motor function acquisitions (head control, sitting and walking) ( P<0.001). In addition, there was a significant difference in the transcription level of fl-SMN2 between the undegenerated group and the degenerated group in sitting and standing ( F=5.432, P=0.023 and F=4.315, P=0.047, respectively). Conclusions:Both the copy number of SMN2 and the transcript level of fl-SMN2 are correlated with SMA severity, survival, and motor milestones, serving as valuable biomarkers for evaluating phenotypic severity of SMA.The transcript level of fl-SMN2 s may play an important role in the degeneration of sitting and standing.

Objective To study the gas analysis of umbilical cord artery blood and radial artery blood on predicating the prognosis of asphyxia neonate?Methods From September 2014 to September 2015, 328 neonates were divided into groups by Apgar score:290 patients in the control group and 27 patients in the mild asphyxia group,11 patients in the severe asphyxia group?After birth,umbilical artery blood,radial artery blood gas analysis was perfomed, oxygenation index was calculated, Outcome of neonatal behavioral neurological assessment ( NBNA) in neonates with asphyxia was regular follow?uped,the relationship between pH value and umbilical artery blood gas analysis was analyzed?Results The pH, PO2, PCO2 and oxygenation index of umbilical cord blood and radial artery blood in the severe asphyxia group was(7?11±0?25,(73?93±23?35) mmHg,(51?36±16?37) mmHg,206?23±98?12),significant different than the mild group(7?24±0?05,(86?35 ±12?56) mmHg,(45?89± 9?21) mmHg,411?22±57?94) and the control group(7?28±0?08,(87?80±12?07) mmHg,(43?68± 6?45) mmHg,426?23±73?30)(P<0?05)?The pH,PO2,PCO2 and oxygenation index of umbilical cord blood and radial artery blood in the severe asphyxia group was(7?25±0?18,(74?66±24?09) mmHg,(51?42±17?83) mmHg,332?03±65?19),significant different than the mild group(7?31±0?09,(87?24 ±11?75) mmHg,(45?73±10?21) mmHg,405?67±82?65) and the control group(7?32±0?06,(87?99±11?81) mmHg,(42?84± 9?32) mmHg,439?89±60?76)(P<0?05)?The NBNA scores of the severe asphyxia group was (34?09±5?02) points,lower than the mild group(36?62±2?04)(F=21?65,P<0?05)?The NBNA scores showed significant relationship with umbilical cord blood pH in the severe asphyxia group( r=0?877,P<0?01)?Conclusion The pH,PO2 and oxygenation index of umbilical cord blood and radial artery blood was lower while PCO2 was markedly high in the severe asphyxia group than other groups?For neonates, there is a correlation between umbilical cord blood pH and NBNAs core, neonates borned with hypoxia and acidosis should monitor blood gas analysis and oxygenation index dynamically

Objective To explore the relative expression of plasma miR-199a-5p and miR-200c-3p in gastric adenocarcinoma cancer(GAC) patients and its clinical value.Methods Case-control study was used in this research.The relative expression of plasma miR-199a-5p and miR-200c-3p from 47 GAC patients and 50 healthy controls were determined by RT-PCR ( TaqMan Probe method).Meanwhile, the association with age, gender, tumor location, size, degree of differentiation, TNM stage, lymph node metastasis and other clinical pathological parameters were analyzed.The expression of these two miRNAs in plasma of 30 GAC patients during preoperation was compared with their expression 6-8 days after radical surgery.The sensitivity and specificity of plasma miRNAs expression for the diagnosis of GAC were analyzed using the receiver operating characteristic ( ROC ) curve.SPSS20.0 statistical software was used for statistical analysis.T-test, paired t-test and one-factor ANOVA were used for normal distribution of quantitative data.Results The plasma level of miR-199a-5p in GAC patients was significantly lower(1.05 ±0.22) (t =3.058,P =0.003), while miR-200c-3p was significantly higher(15.15 ±3.02) (t =-2.854,P=0.006), when they were compared with those in controls(26.80 ±8.38, 3.39 ±0.87).Low miR-199a-5p expression in GAC patients were associated with lymph node metastasis ( F =4.725, P =0.029) and the differentiation degree of gastric cancer(F=3.854,P=0.032).The relative expression of miR-199a-5p in postoperative plasma was significantly increased(t=-3.814,P=0.001), but the relative expression of miR-200c-3p was significantly reduced when compared to the preoperative samples(t=2.978, P=0.006).Area under the ROC curve of miR-199a-5p, miR-200c-3p and combined miR-199a-5p and miR-200c-3p were 0.692, 0.792 and 0.798, the sensitivity and specificity were 87%,97%,92.5% and 43%,54%, 65%, respectively.Conclusion Combined detection of miR-199a-5p and miR-200c-3p in plasma has a higher sensitivity and specificity than the conventional tumor marker CEA and CA19-9, and may be a useful combination for gastric cancer diagnosis.

Objective To systematically review the diagnostic value of microRNA(miRNA) quantitation in breast cancer .Meth-ods Literatures about miRNA and breast cancer diagnosis were selected by retrieving Medline ,Embase and Cochrane Library .Ac-cording to the inclusion and exclusion criteria ,the literatures were independently screened ,and a 2 × 2 contingency table was con-structed .Quality of literatures was assessed by quality assessment of diagnostic accuracy studies(QUADAS) .Statistical analysis was performed by employing Meta-Disc 1 .4 software and STATA 11 .0 .Results 16 studies were included ,which contained 1 303 patients and 711 control samples .There were threshold effects among these studies (the spearman′s correlation coefficient was-0 .758 ,P=0 .001) .A random effects model was used for meta-analysis .The summary sensitivity ,specificity ,positive likelihood ratio ,negative likelihood ratio ,and diagnostic odds ratio for miRNA in breast cancer diagnosis were 0 .77(95% CI:0 .75 -0 .79) , 0 .77(95% CI:0 .74-0 .80) ,4 .19(95% CI:2 .79-6 .30) ,0 .25(95% CI:0 .19-0 .35) ,19 .91(95% CI:9 .68-40 .95) .The area un-der curve of SROC was 0 .895 0 .Conclusion These results suggest that miRNAs have potential value to diagnose breast cancer . However ,effective diagnosis of breast cancer still needs to be conducted with assistance of clinical findings and traditional lab inves-tigations .