Guided by the concept of quality by design(QbD), this study optimizes the calcination and quenching process of calcined Magnetitum and establishes the XRD characteristic spectra of calcined Magnetitum, providing a scientific basis for the formulation of quality standards. Based on the processing methods and quality requirements of Magnetitum in the Chinese Pharmacopoeia, the critical process parameters(CPPs) identified were calcination temperature, calcination time, particle size, laying thickness, and the number of vinegar quenching cycles. The critical quality attributes(CQAs) included Fe mass fraction, Fe~(2+) dissolution, and surface color. The weight coefficients were determined by combining Analytic Hierarchy Process(AHP) and the criteria importance though intercrieria correlation(CRITIC) method, and the calcination process was optimized using orthogonal experimentation. Surface color was selected as a CQA, and based on the principle of color value, the surface color of calcined Magnetitum was objectively quantified. The vinegar quenching process was then optimized to determine the best processing conditions. X-ray diffraction(XRD) was used to establish the characteristic spectra of calcined Magnetitum, and methods such as similarity evaluation, cluster analysis, and orthogonal partial least squares-discriminant analysis(OPLS-DA) were used to evaluate the quality of the spectra. The optimized calcined Magnetitum preparation process was found to be calcination at 750 ℃ for 1 h, with a laying thickness of 4 cm, a particle size of 0.4-0.8 cm, and one vinegar quenching cycle(Magnetitum-vinegar ratio 10∶3), which was stable and feasible. The XRD characteristic spectra analysis method, featuring 9 common peaks as fingerprint information, was established. The average correlation coefficient ranged from 0.839 5-0.988 1, and the average angle cosine ranged from 0.914 4 to 0.995 6, indicating good similarity. Cluster analysis results showed that Magnetitum and calcined Magnetitum could be grouped together, with similar compositions. OPLS-DA discriminant analysis identified three key characteristic peaks, with Fe_2O_3 being the distinguishing component between the two. The final optimized processing method is stable and feasible, and the XRD characteristic spectra of calcined Magnetitum was initially established, providing a reference for subsequent quality control and the formulation of quality standards for calcined Magnetitum.
X-Ray Diffraction/methods* ; Drugs, Chinese Herbal/chemistry* ; Quality Control ; Particle Size

This study aims to investigate the effect of Chaijin Jieyu Anshen Formula on the neuroinflammation of rats with insomnia complicated with depression through the regulation of triggering receptor expressed on myeloid cells 2(TREM2)/complement protein C1q signaling pathway. Rats were randomly divided into a normal group, a model group, a positive drug group, as well as a high, medium, and low-dose groups of Chaijin Jieyu Anshen Formula, with 10 rats in each group. Except for the normal group, the other groups were injected with p-chlorophenylalanine and exposed to chronic unpredictable mild stress to establish the rat model of insomnia complicated with depression. The sucrose preference experiment, open field experiment, and water maze test were performed to evaluate the depression in rats. Enzyme-linked immunosorbent assay was employed to detect serum 5-hydroxytryptamine(5-HT), dopamine(DA), and norepinephrine(NE) levels. Hematoxylin and eosin staining and Nissl staining were used to observe the damage in nucleus accumbens neurons. Western blot and immunofluorescence were performed to detect TREM2, C1q, postsynaptic density 95(PSD-95), and synaptophysin 1(SYN1) expressions in rat nucleus accumbens, respectively. Golgi-Cox staining was utilized to observe the synaptic spine density of nucleus accumbens neurons. The results show that, compared with the model group, Chaijin Jieyu Anshen Formula can significantly increase the sucrose preference as well as the distance and number of voluntary activities, shorten the immobility time in forced swimming test and the successful incubation period of positioning navigation, and prolong the stay time of space exploration in the target quadrant test. The serum 5-HT, DA, and NE contents in the model group are significantly lower than those in the normal group, with the above contents significantly increased after the intervention of Chaijin Jieyu Anshen Formula. In addition, Chaijin Jieyu Anshen Formula can alleviate pathological damages such as swelling and loose arrangement of tissue cells in the nucleus accumbens, while increasing the Nissl body numbers. Chaijin Jieyu Anshen Formula can improve synaptic damage in the nucleus accumbens and increase the synaptic spine density. Compared to the normal group, the expression of C1q protein was significantly higher in the model group, while the expression of TREM2 protein was significantly lower. Compared to the model group, the intervention with Chaijin Jieyu Anshen Formula significantly downregulated the expression of C1q protein and significantly upregulated the expression of TREM2. Compared with the model group, the PSD-95 and SYN1 fluorescence intensity is significantly increased in the groups receiving different doses of Chaijin Jieyu Anshen Formula. In summary, Chaijin Jieyu Anshen Formula can reduce the C1q protein expression, relieve the TREM2 inhibition, and promote the synapse-related proteins PSD-95 and SNY1 expression. Chaijin Jieyu Anshen Formula improves synaptic injury of the nucleus accumbens neurons, thereby treating insomnia complicated with depression.
Animals ; Male ; Rats ; Nucleus Accumbens/metabolism* ; Drugs, Chinese Herbal/administration & dosage* ; Depression/complications* ; Membrane Glycoproteins/genetics* ; Rats, Sprague-Dawley ; Sleep Initiation and Maintenance Disorders/complications* ; Neurons/metabolism* ; Receptors, Immunologic/genetics* ; Signal Transduction/drug effects* ; Synapses/metabolism*

Stem cells play a crucial role in maintaining tissue regenerative capacity and homeostasis. However, mechanisms associated with stem cell senescence require further investigation. In this study, we conducted a proteomic analysis of human dental pulp stem cells (HDPSCs) obtained from individuals of various ages. Our findings showed that the expression of NUP62 was decreased in aged HDPSCs. We discovered that NUP62 alleviated senescence-associated phenotypes and enhanced differentiation potential both in vitro and in vivo. Conversely, the knocking down of NUP62 expression aggravated the senescence-associated phenotypes and impaired the proliferation and migration capacity of HDPSCs. Through RNA-sequence and decoding the epigenomic landscapes remodeled induced by NUP62 overexpression, we found that NUP62 helps alleviate senescence in HDPSCs by enhancing the nuclear transport of the transcription factor E2F1. This, in turn, stimulates the transcription of the epigenetic enzyme NSD2. Finally, the overexpression of NUP62 influences the H3K36me2 and H3K36me3 modifications of anti-aging genes (HMGA1, HMGA2, and SIRT6). Our results demonstrated that NUP62 regulates the fate of HDPSCs via NSD2-dependent epigenetic reprogramming.
Humans ; Dental Pulp/cytology* ; Nuclear Pore Complex Proteins/genetics* ; Cellular Senescence/genetics* ; Stem Cells/metabolism* ; Epigenesis, Genetic ; Cell Proliferation ; Cell Differentiation ; Histone-Lysine N-Methyltransferase/metabolism* ; Cells, Cultured ; Cellular Reprogramming ; Cell Movement ; Proteomics

Objective To compare the immediate effects of acupuncture at the true and false acupoints of Yanglingquan on functional connectivity in sensorimotor network(SMN)and dorsal attentional network(DAN)of stroke patients based on functional magnetic resonance imaging(fMRI)technology;To explore the central regulatory mechanism and acupoint specificity of acupuncture in stroke patients with hemiplegia.Methods Totally 20 patients with stroke and hemiplegia were included in the study.fMRI scans of acupuncture at the true and false acupoints of Yanglingquan were performed once every 2 weeks,and motion-related SMN and DAN were extracted by independent component analysis to compare the differences in functional connectivity.Results In SMN,after acupuncture at the Yanglingquan true acupoint,the functional connectivity was enhanced compared with before acupuncture.The enhanced brain areas included the right anterior central gyrus,superior temporal gyrus,inferior frontal gyrus,cuneiform lobe,and anterior cuneiform lobe,as well as the left middle temporal gyrus,occipital gyrus,superior temporal gyrus,parahippocampal gyrus,inferior frontal gyrus,and superior temporal gyrus.After acupuncture at the Yanglingquan false acupoint,the functional connectivity was enhanced compared with before acupuncture.The enhanced brain areas included the right anterior central gyrus,superior frontal gyrus,middle frontal gyrus,and cingulate gyrus,as well as the left medial frontal gyrus,anterior cingulate gyrus,lentiform nucleus,and caudate nucleus.In DAN,after acupuncture at the Yanglingquan true acupoint,the functional connectivity was enhanced compared with before acupuncture.The enhanced brain areas included the right anterior cingulate lobe,superior temporal gyrus,middle temporal gyrus,and occipital gyrus,as well as the left cingulate gyrus,posterior cingulate gyrus,and anterior cingulate lobe.After acupuncture at the Yanglingquan false acupoint,the functional connectivity was enhanced compared with before acupuncture,and the enhanced brain areas included the right anterior cingulate gyrus,left anterior cingulate gyrus,and medial frontal gyrus.Conclusion Acupuncture at Yanglingquan can activate SMN and DAN bilateral related brain regions in patients with hemiplegia,which may promote the recovery of motor function by regulating the initiation and execution of motor activities,and has more acupoint specificity compared with false acupoint.

Aim To explore the effect of total flavonoid extract(TFE)of Ampelopsis megalophylla on the pro-liferation and apoptosis of human breast cancer cells and its mechanism in autophagy inhibition.Methods For human cervical cancer cell Hela,human lung cancer cell A549,human liver cancer cell SMMC-7721,human breast cancer cell MCF-7,MDA-MB-231 and human normal liver cell L-02,MTT method was used to select sensitive cell lines.The inhibitory effect of TFE combined with autophagy inhibitor 3-methylade-nine(3-MA)on sensitive cell proliferation was detec-ted using MTT assay.The morphological changes of cells were observed using transmission electron micros-copy and Hoechst 33258 single staining method.The changes in cell apoptosis rate were detected using An-nexin V-FITC/PI dual staining method.The expression levels of apoptosis related proteins and pathway pro-teins(death receptor pathway,mitochondrial pathway,endoplasmic reticulum stress pathway)were detected uisng Western blot.The expression of the key protein Cyt-c in mitochondrial pathway was determiend by im-munofluorescence,and the autophagy agonist rapamy-cin was selected for reverse validation.Results TFE could inhibit the proliferation of human breast cancer cells in a concentration-dependent manner,and MCF-7 cells were sensitive cell lines.Compared with the TFE group,the TFE+3-MA group significantly increased the inhibition rate of MCF-7 cells at 24,48,and 72 h(P＜0.01).The number of cells decreased,the gap increased,the number of apoptotic bodies increased,and the apoptosis rate increased(P＜0.01).The ex-pression levels of Bax/Bcl-2(P＜0.01),cleaved-caspase3(P＜0.01),Cyt-c(P＜0.05),FADD,and cleaved-caspase-12 all increased,and the expres-sion of apoptotic protein Cyt-c in nucleus increased.The fluorescence of the TFE+RA group decreased,re-versing the mitochondrial pathway apoptosis induced by TFE.Conclusions TFE can significantly inhibit the proliferation of human breast cancer cells.When inhib-iting autophagy,it may promote the apoptosis of MCF-7 cells through the mitochondrial pathway,and activa-ting autophagy can reverse apoptosis.

Objective To establish a high-throughput screening system to obtain key Staphylococcus aureus (S.aureus)secretory proteins which required for S.aureus survival in macrophages.Methods Based on our validated eukaryotic expression vector library of S.aureus secretory proteins,DNA transfection was used to obtain an RAW264.7 macrophage array expressing S.aureus secretory proteins.After the RAW264.7 cells were infected with S.aureus,the extracellular bacteria were removed to observe the intracellular surviving situation of S.aureus.Finally,the screening results were validated by the overexpression and knockout S.aureus of corresponding secretory proteins.Results The optimal transfection dose (1.0 μg/well)of plasmids for RAW264.7,multiplicity of infection (MOI,1 .0 ),and infection time (4 h after removing extracellular bacteria of S.aureus ) were established respectively.To validate the screening results,the corresponding overexpression and knockout strains were constructed.And hypothetical protein and Serine protease E were found to promote the survival of intracellular S.aureus.Conclusion We successfully construct a screening system for key secreted secretory proteins which required for S.aureus surviving in macrophages,which may advance the study of the intracellular surviving mechanism of S.aureus.

Abstract: Objective　To investigate the antimicrobial activity of omadacycline (OMC) against clinical Streptococcus agalactiae (GBS) isolates, as well as its relationship with biofilm formation, resistance genes and virulence genes. Methods　A total of 136 strains of Streptococcus agalactiae isolated from Shenzhen Nanshan People's Hospital between 2015 to 2020. The minimum inhibitory concentration (MIC) of OMC against Streptococcus agalactiae was determined by broth microdilution. Crystal violet staining was used to detect the biofilm formation ability of GBS. Resistance genes (tetM, tetO, tetK, ermB, OptrA) and virulence genes (cpsⅢ, bca, fbsA, cpsA, scpB) were investigated by polymerase chain reaction (PCR). Results　Among the 136 clinical isolates of GBS, 20 strains (14.7%) were resistant to OMC, 64 (47.1%) were intermediate, and 52 (38.2%) were sensitive. Fifty-seven strains (41.9%) were biofilm-positive, 20 of which (35.1%) were sensitive to OMC. Seventy-nine strains (58.1%) were biofilm-negative, 32 of which (40.5%) were susceptible to OMC. There was a statistically significant difference in the sensitivity rates between the two groups of strains (χ2=63.062, P<0.001), but there was no significant difference in the sensitivity of OMC among the biofilm-positive strains (Fisher's exact test, P=0.824). The resistance rates of tetM, tetO, ermB and OptrA positive strains were higher than those of negative strains, while tetK was opposite. The presence of tetM (Z=0.815, P=0.415), tetO (Z=0.151, P=0.88), tetK (Z=0.567, P=0.571), ermB (Z=1.198, P=0.231) resistance genes in Streptococcus agalactiae had no significant impact on the sensitivity of OMC. However, the presence of the OptrA resistance gene showed a statistically significant effect on the sensitivity of OMC (Z=2.913, P=0.004). The virulence factors cpsⅢ, bca, fbsA, cpsA and scpB were all detected at a rate higher than 50%. The presence of the virulence genes cpsⅢ (Z=0.222, P=0.824), bca (Z=0.141, P=0.888), fbsA (Z=0.813, P=0.416), and cpsA (Z=1.615, P=0.106) in Streptococcus agalactiae had no significant impact on the sensitivity of OMC. However, there was a significant inter-group difference in the scpB virulence gene (Z=2.844, P=0.004), but the rank mean values and resistance rates of scpB-positive strains were lower than those of the negative strains. Conclusions　The formation of biofilm in Streptococcus agalactiae reduces its sensitivity to OMC, but there was no significant difference in the sensitivity to OMC among the biofilm-positive strains. The presence of resistance genes tetM, tetO, tetK, ermB, and virulence genes cpsⅢ, bca, fbsA, cpsA, scpB in Streptococcus agalactiae is not associated with OMC resistance, but the presence of the resistance gene OptrA is correlated with OMC resistance..

OBJECTIVE: To investigate CT values of cancellous bone in femoral neck in adults over 60 years with proximal femoral fractures. METHODS: From January 2020 to December 2020, a retrospective analysis was performed on 280 subjects aged 60 years or older who underwent bilateral hip CT examination, including 85 males and 195 females, 120 on the left side and 160 on the right side, aged 75 (66, 82) years old. One hundred thirty-six patients with proximal femoral fractures were included in study group and 144 patients without fractures were included in control group. GEOptima CT was used to scan and reconstruct horizontal, coronal and sagittal layers of proximal femur. CT values of cancellous bone in femoral neck were measured and compared between two groups. The relationship between CT values of cancellous bone of femoral neck and proximal femoral fracture was analyzed statistically. RESULTS: In terms of age, fracture group aged 79(73.3, 85.0) years old, non-fracture group aged 69.5 (64.0, 78.8) years old, and had significant difference in age between two groups (P<0.05). In terms of CT value, regional CT value in fracture group was 8.62(-3.62, 27.15) HU, which was lower than that in non-fracture group 34.31(-5.93, 71.74) HU(P<0.05). CT value on coronal view in fracture group was -8.48(-30.96, 17.46) HU, which was lower than that in non-fracture group 40.49(5.55, 80.71) HU (P<0.05). CT value on sagittal view in fracture group was -31.28(-54.91, -5.11) HU, which was lower than that in non-fracture group 7.74(-20.12, 44.54) HU (P<0.05). CT values on horizontal view in fracture group was 0.17(-23.13, 24.60) HU, which was lower than that in non-fracture group 46.40(10.42, 85.18) HU(P<0.05). The mean regional CT values among three planes in the fracture group were lower than those in the non-fracture group. Logistic regression analysis showed coronal CT value was influencing factors of proximal femoral fracture, and it could be written into regression equations that predict probability of fracture. CONCLUSION In adults aged over 60 years old, CT values of cancellous bone of femoral neck decreased with increasing age. The smaller CT value of cancellous bone of femoral neck, the greater risk of proximal femoral fracture.
Male ; Adult ; Female ; Humans ; Middle Aged ; Aged ; Aged, 80 and over ; Proximal Femoral Fractures ; Retrospective Studies ; Femur Neck ; Hip Fractures/surgery* ; Tomography, X-Ray Computed ; Bone Density

PURPOSE: As common clinical screening tests cannot effectively predict a difficult airway, and unanticipated difficult laryngoscopy remains a challenge for physicians. We herein used ultrasound to develop some point-of-care predictors for difficult laryngoscopy. METHODS: This prospective observational study included 502 patients who underwent laryngoscopy and a detailed sonographic assessment. Patients under 18 years old, or with maxillofacial deformities or fractures, limited mouth opening, limited neck movement or history of neck surgery were excluded from the study. Laryngoscopic views of all patients were scored and grouping using the modified Cormack-Lehane (CL) scoring system. The measurements acquired comprised tongue width, the longitudinal cross-sectional area of the tongue, tongue volume, the mandible-hyoid bone distance, the hyoid bone-glottis distance, the mandible-hyoid bone-glottis angle, the skin-thyrohyoid membrane distance, the glottis-superior edge of the thyroid cartilage distance (DGTC), the skin-hyoid bone distance, and the epiglottis midway-skin distance. ANOVA and Chi-square were used to compare differences between groups. Logistic regression was used to identify risk factors for difficult laryngoscopy and it was visualized by receiver operating characteristic curves and nomogram. R version 3.6.3 and SPSS version 26.0 were used for statistical analyses. RESULTS: Difficult laryngoscopy was indicated in 49 patients (CL grade Ⅲ - Ⅳ) and easy laryngoscopy in 453 patients (CL grade Ⅰ - Ⅱ). The ultrasound-measured mandible-hyoid bone-glottis angle and DGTC significantly differed between the 2 groups (p < 0.001). Difficult laryngoscopy was predicted by an area under the curve (AUC) of 0.930 with a threshold mandible-hyoid bone-glottis angle of 125.5° and by an AUC of 0.722 with a threshold DGTC of 1.22 cm. The longitudinal cross-sectional area of the tongue, tongue width, tongue volume, the mandible-hyoid distance, and the hyoid-glottis distance did not significantly differ between the groups. CONCLUSION Difficult laryngoscopy may be anticipated in patients in whom the mandible-hyoid bone-glottis angle is smaller than 125.5° or DGTC is larger than 1.22 cm.
Humans ; Adolescent ; Laryngoscopy ; Prospective Studies ; Tongue/diagnostic imaging* ; Respiratory System ; Ultrasonography

It has been well documented that exercise can improve bone metabolism, promote bone growth and development, and alleviate bone loss. MicroRNAs (miRNAs) are widely involved in the proliferation and differentiation of bone marrow mesenchymal stem cells, osteoblasts, osteoclasts and other bone tissue cells, and regulation of balance between bone formation and bone resorption by targeting osteogenic factors or bone resorption factors. Thus miRNAs play an important role in the regulation of bone metabolism. Recently, regulation of miRNAs are shown to be one of the ways by which exercise or mechanical stress promotes the positive balance of bone metabolism. Exercise induces changes of miRNAs expression in bone tissue and regulates the expression of related osteogenic factors or bone resorption factors, to further strengthen the osteogenic effect of exercise. This review summarizes relevant studies on the mechanism whereby exercise regulates bone metabolism via miRNAs, providing a theoretical basis for osteoporosis prevention and treatment with exercise.
Humans ; MicroRNAs/metabolism* ; Osteogenesis/genetics* ; Cell Differentiation ; Osteoblasts ; Bone Resorption/metabolism*