ObjectiveTo study the synergistic effect of p38MAPK and ERK1/2 in bone marrow mesenchymal stem cells (BMSCs), and to explore their influence on osteogenic differentiation in BMSCs cultures. MethodsMouse BMSCs were cultured in phenol red-free α-MEM containing osteogenic supplements (OS) for inducing osteogenic differentiation. The temporal sequence of osteogenic differentiation in BMSCs cultures was assayed by measuring alkaline phosphatase activity (ALP) and calcium deposition gene expression. The activation of p38MAPK and ERK1/2 was detected by western blotting using phospho-specific MAP kinase antibody. BMSCs were treated with the inhibitor of p38MAPK pathway(SB203580) or ERK1/2 pathway (PD98059), and osteogenic differentiation was measured. BMSCs were treated with SB203580 or sodium arsenite(ARS), a strong activator of p38MAPK, and the phosphorylation of ERK 1/2 was measured. BMSCs were treated with PP2A inhibitor, Okadaic acid(OA), the phosphorylation of ERK1/2 and osteogenic differentiation were measured. lmmunoprecipitation was used to test the binding interaction between PP2A and ERK1/2, and the effect of SB203580 on the interaction. ResultsTreatment of BMSCs with osteogenic supplements resulted in activation of p38MAPK and ERK1/2 that coincided with osteogenic differentiation. Inhibition of p38MAPK activation by SB203580, blocked the osteogenic differentiation, whereas inhibition of ERK1/2 activation by PD98059, enhanced the osteogenic differentiation in a dose-dependent manner.SB203580 treatment resulted in increased ERK1/2 phosphorylation. By contrast, ARS treatment resulted in decreased ERK1/2 phosphorylation. Inhibition of PP2A by OA resulted in increased ERK1/2 phosphorylation. OS-induced osteogenic differentiation was also attenuated by PP2A inhibition. Immunoprecipitation confirmed the association of PP2A with ERK1/2 in BMSCs cultures, which was decreased by SB203580 treatment. ConclusionThe present study demonstrates a synergistic effect between p38MAPK and ERK1/2 signaling pathways via PP2A in BMSCs cultures, which may regulate the osteogenic differentiation of BMSCs.

Objective:To evaluate the effect of micro-implant assisted rapid palatal expansion (MARPE) in patients with different radiographic stages of midpalatal suture maturation.Methods:Twenty-eight patients [7 males and 21 females; age (15.5±5.5) years] with maxillary transverse deficiency were selected in the Department of Orthodontics, Hospital of Stomatology, Jilin University from February 2017 to October 2019. According to the fusion of the midpalatal suture, the patients were divided into two groups: 10 patients [3 males and 7 females; age (19.9±6.9) years] were in fusion group and 18 patients [4 males and 14 females; age (13.0±2.4) years] were in non-fusion group. Each patient had cone-beam CT (CBCT) images taken immediately after the placement of micro-implants (T1) and the completion of maxillary expansion (T2). The CBCT images were analyzed using the Dolphin Imaging software to evaluate the amount of midpalatal expansion and the percentages of bony expansion, etc. Statistical analysis was carried out to compare the two groups.Results:After MARPE treatment, the amount of sutural expansion in fusion group was (3.23±1.45) mm while that in non-fusion group was (4.97±1.47) mm ( P<0.05). There was a statistically significant difference in the percentages of bony expansion efficiency between the non-fusion group (71±20)% and the fusion group (46±18)% ( P<0.05). Conclusions:Bony expansion efficiency was affacted by the radiographic stages of midpalatal suture maturation.

Objective To analyze the main ingredients and investigate the anti-psoriasis effect in the standard decoction of Lithospermum. Methods The extraction rate was determined by extract determination method, and the content of total polysaccharide and total phenolic acid was evaluated by spectrophotometry. Moreover, the effects of different concentrations of the standard decoction of Lithospermum on skin lesions induced by imiquimod (IMQ) in psoriatic mice were observed. Results The extracting rate was 4.65 ％, the total polysaccharide content was 1.182 mg/ml and the total phenolic acid content was 3.506 mg/ml. The different concentrations of the standard decoction of Lithospermum could ameliorate the scales, erythema and psoriasis-like mice skin and reduce the thickness of epidermis. Conclusions The standard decoction of Lithospermum could improve the psoriasis-like lesions induced by imiquimod in mice and possess the anti-psoriasis effect.

Objective:To investigate the effects of medical maggot excretions/secretions (ES) on neutrophils phagocytosis and bactericidal effect in patients with diabetic foot ulcer (DFU).Methods:The experimental research method was used. Thirty DFU patients (16 males and 14 females, aged (64±7) years)who were admitted to the Diabetes Foot Center, the Department of Endocrinology of Air Force Hospital of Eastern Theater Command from June to December 2020 and met the inclusion criteria were recruited. Discontinuous percoll gradient centrifugation method was used to separate the neutrophils. Cells from each patient were enrolled into normal saline group and maggot ES group (30 wells in each group), respectively; sterile normal saline and ES with a final mass concentration of 357 μg/mL (the same as below) were added, respectively. After 1 and 2 hour(s) of culture, the phagocytosis rate and phagocytic index of cells were observed and counted under Wright's staining. Ten patients were selected, then the cells of each patient were enrolled into Pseudomonas aeruginosa+neutrophils group and Pseudomonas aeruginosa+neutrophils+maggot ES group (10 wells in each group) and were treated corresponding, respectively. Pseudomonas aeruginosa alone group and Pseudomonas aeruginosa+maggot ES group (10 wells in each group) were set up respectively; Pseudomonas aeruginosa+RPMI 1640 culture medium+sterile normal saline and Pseudomonas aeruginosa+RPMI 1640 culture medium+maggot ES were added, respectively. After 2 hours of culture, the number of viable bacteria colony was counted by plate colony number method. Six, six, and three patients were selected respectively, and the cells of each patient were respectively enrolled into maggot ES group and normal saline group (6, 6, and 3 wells in each group, respectively) and treated accordingly. After 6 hours of culture, real-time fluorescent quantitative reverse transcription polymerase chain reaction was used to detect the mRNA expressions of interleukin 1β (IL-1β), IL-6, and lysozyme in cells, the content of IL-1β and IL-6 in cell culture supernatant were determined by enzyme-linked immunosorbent assay, and the positive cells expressing lysozyme were observed with immunofluorescence method. Data were statistically analyzed with one-way analysis of variance, paired sample t test, least significant difference test, and Wilcoxon rank sum test. Results:After 1 hour of culture, the phagocytosis rate and phagocytic index of cells in maggot ES group (53.5% (49.7%, 58.0%) and 3.18 (2.96, 3.32)) were similar to 52.0% (47.5%, 55.2%) and 3.15 (2.96, 3.25) of normal saline group ( Z=-1.701, -1.092, P>0.05). After 2 hours of culture, the phagocytosis rate and phagocytic index of cells in maggot ES group (70.0% (66.7%, 72.0%) and 4.47 (4.22, 4.96)) were significantly higher than 58.0% (55.0%, 60.0%) and 4.11 (3.52, 4.24) in normal saline group ( Z=-4.786, -4.279, P<0.01). After 2 hours of culture, the number of viable bacteria colony in Pseudomonas aeruginosa+neutrophils group was significantly lower than that in Pseudomonas aeruginosa alone group ( P<0.01), and the number of viable bacteria colony in Pseudomonas aeruginosa+neutrophils+maggot ES group was significantly lower than that in Pseudomonas aeruginosa+maggot ES group and Pseudomonas aeruginosa+neutrophils group ( P<0.01). After 6 hours of culture, the mRNA expressions of IL-1β, IL-6, and lysozyme of cells in maggot ES group were significantly higher those in normal saline group ( t=-3.279, -4.273, -4.763, P<0.05 or P<0.01); the concent of IL-1β and IL-6 in cell culture supernatant of maggot ES group were significantly higher than those of normal saline group ( t=-9.526, -6.447, P<0.01); there were significantly more positive cells expressing lysozyme in maggot ES group than in normal saline group. Conclusions:Maggot ES can enhance the phagocytosis and bactericidal effect of neutrophils on Pseudomonas aeruginosa by promoting the production of neutrophils immune defense related cytokines and lysozyme in DFU patients.

BACKGROUND:Osteonecrosis of the femoral head is a common and disabling disease,which is mainly characterized by microcirculation disorders and bone cell metabolism disorders.Luzhongjiangu decoction was developed by Shandong Academy of Chinese Medicine and used in the form of soup in the clinic,which has good efficacy in the treatment of osteonecrosis of the femoral head.However,its mechanism of action has not been clarified. OBJECTIVE:To study the effect mechanism of Luzhongjiangu decoction on intestinal flora in rats with osteonecrosis of the femoral head based on 16S rDNA sequencing technique. METHODS:The model of osteonecrosis of the femoral head was established in Wistar rats by intragastric administration of retinoic acid.The therapeutic effect of Luzhongjiangu decoction was evaluated by serum hormone,bone histopathology and serum hormone levels.16s rDNA sequencing technique was used to detect the intestinal flora of rats in the blank control group,model group and middle-dose Luzhongjiangu decoction group.The corresponding library was constructed and OTU clustering and microbial community diversity and abundance analysis were carried out to determine the composition of intestinal flora and the changes of species and diversity among groups. RESULTS AND CONCLUSION:Luzhongjiangu decoction could significantly increase the expression of osteocalcin,osteopontin and other osteogenic related factors,alleviate the destruction of bone trabeculae,increase bone mineral density,and had a significant therapeutic effect on osteonecrosis of the femoral head,of which the middle dose group showed the most significant effect.The results of intestinal flora sequencing showed that Luzhongjiangu decoction improved the flora disorder of rats with osteonecrosis of the femoral head to some extent,and screened out different colonies such as Bacillus,Desulfurizans,Desulfurization,Isobacteria,Bifidobacterium and so on;it could up-regulate the abundance of beneficial bacteria such as Bifidobacterium,down-regulate the abundance of harmful bacteria such as Desulfovibrio,and improve the structure of intestinal flora.Functional prediction analysis indicated that Luzhongjiangu decoction could mainly affect amino acid metabolism and energy metabolism.Correlation analysis showed that the differential bacteria of Bifidobacterium and Intestinimonas in the middle dose group of Luzhongjiangu decoction were positively correlated with vitamin D3,estradiol and calcitonin,and negatively correlated with prostaglandin E2.In the model group,Escherichia-Shigella,Desulfovibrio,Globicatella and Streptococcus were positively correlated with prostaglandin E2 and negatively correlated with vitamin D3,estradiol and calcitonin.To conclude,Luzhongjiangu decoction may play a role in the treatment of osteonecrosis of the femoral head by regulating the structure of intestinal flora,up-regulating the abundance of beneficial bacteria and affecting the secretion of vitamin D3,estradiol,calcitonin and prostaglandin E2.

OBJECTIVE To optimize the freeze-drying process for sheep placenta slices. METHODS An orthogonal test design was used with pre-freezing time， drying time and drying temperature as indicators to screen for the optimal freeze-drying process for sheep placenta slices. The peptide content， ethanol-soluble extract content， and freeze-drying rate of sheep placenta were used as indicators，the analytic hierarchy process-criteria importance through intercriteria correlation （AHP-CRITIC） method was employed to determine the weight of each indicator and calculate the comprehensive score， which was verified using the technique for order of preference by similarity to ideal solution （TOPSIS） model. RESULTS The optimal preparation process was found to be the pre-freezing time of 2 hours， the drying time of 16 hours， and the drying temperature of 30 °C. The average values of peptide content， ethanol-soluble extract content， and freeze-drying rate for three batches of samples were 5.883 mg/mL， 27.1%， and 95.77%， respectively； the comprehensive scores of three batches were 96.42， 99.18 and 99.58， with RSD of 1.75%. CONCLUSIONS This study successfully optimized the freeze-drying process for sheep placenta slices， which can provide a reference for the quality standard setting and industrial production of this type of slice.

BACKGROUND:Our previous studies found that the polypeptide of Cornus cervi Colla can promote bone growth,which has a good application prospect in the treatment of bone diseases.However,how Cornus cervi Colla works in the body and the principle are not clear. OBJECTIVE:To study the in vivo distribution and tracing of Cornus cervi Colla using fluorescence labeling and tracer technique. METHODS:Cornus cervi Colla was fluorescently labeled using fluorescein isothiocyanate,and the labeling results were detected by fluorescence imaging and UV spectral scanning.Successfully labeled Cornus cervi Colla was injected into mice by gavage,and the absorption of Cornus cervi Colla into blood was detected by laser confocal microscopy,and the distribution of Cornus cervi Colla in mice was detected by small animal in vivo imager.The distribution of Cornus cervi Colla in the mice was detected by laser confocal microscopy.Samples were taken from serum and bone at the time of the strongest fluorescence,and gel electrophoresis was carried out on serum and bone tissue protein solutions,and the components of Cornus cervi Colla absorbed into target organs were determined by secondary mass spectrometry. RESULTS AND CONCLUSION:The fluorescent markers were successfully separated by dextran gel chromatography,and the fluorescence imaging and ultraviolet spectrum scanning proved that the labeling was successful,and the fluorescence substitution degree of FITC-labeled Cornus cervi Colla was 0.953%.The fluorescence intensity of the components of Cornus cervi Colla in the blood showed that Cornus cervi Colla was most distributed in serum after oral administration for 2 hours.The fluorescence images of mice at different times were the same as those of bilateral femur and tibia,indicating that Cornus cervi Colla could play a role by entering the bone.Compared with UniProt database,secondary mass spectrometry showed that the peptide was a characteristic fragment of decorin.It is proved that decorin in Cornus cervi Colla can enter the bone to play a therapeutic role.

Objective To systematically evaluate the efficacy of Xixian Tongshuan Capsules/Pills in the treatment of acute ischemic stroke(AIS).Methods Literature about Xixian Tongshuan Preparation combined with conventional Western medicine for the treatment of AIS was retrieved from CNKI,SinoMed,VIP,Wanfang Data,PubMed,Medline,Embase,Cochrane Library and Web of Science from establishment of the databases to February 28,2023.Meta-analysis was conducted for the studies that could be quantitatively analyzed.The effective rate and response indicators were combined.Results A total of 7 articles were included for Meta-analysis.Results showed that there was statistical difference in the effective rate(RR=0.34,95%CI[0.23,0.51],P<0.01),NIHSS score(MD=-2.90,95%CI[-3.74,-2.06],P<0.01),BI score(MD=-10.08,95%CI[-13.47,-6.68],P<0.01),FIB(MD=-1.18,95%CI[-1.59,-0.77],P<0.01)of Xixian Tongshuan Preparation combined with conventional Western medicine for the treatment of AIS.There was no statistical difference in IL-6(MD=-15.4,95%CI[-33.3,2.49],P=0.09).There was no statistical difference in the effects of different dosage forms and treatment courses on the effective rate and NIHSS score.Conclusion The combination of Xixian Tongshuan Capsules/Pills could better improve the NIHSS and BI scores of patients with AIS,recovery the neurological function,and reduce the risk of blood hypercoagulability by reducing FIB content,with good safety.

Objective To systematically evaluate the efficacy and safety of compound Duzhong Jiangu granules in the treatment of knee osteoarthritis.Methods According to PICOS principle,literature retrieval was carried out in CNKI,SinoMed,VIP,WanFang,PubMed,Medline,Embase and Cochrane Library,and the retrieval time was up to October 2023.Meta-analysis was performed on quan-tifiable studies,and the effective rate and efficacy indicators were combined.Results A total of 5 articles were included for Meta-anal-ysis.The effective rate of compound guzhong Jiangu granules was higher than that of control group,the difference was statistically signifi-cant(RR=1.12,95％CI:1.07-1.18,P＜0.001).Compared with the control group,there were significant differences in pain or discomfort scores during bed rest at night,standing from seat score,maximum walking distance score,getting off standard ladder score,walking on uneven road score and WOMAC score between compound duzhong jiangu granules and the control group(P＜0.01).There were no significant differences in improving morning stiffness or pain after getting up,boarding standard ladder score and joint function ef-ficacy score between compound Duzhong Jiangu Granules and control group(P＞0.05).There were no serious adverse events were repor-ted in the included studies.Conclusion Combined with compound guzhong jiangu granules can improve the therapeutic effect of knee os-teoarthritis,relieve a mild to moderate pain to a certain extent,and promote the recovery of joint function,but it is not good for improving morning stiffness,so it is recommended for early combined treatment with good safety.

OBJECTIVE To establish a quantitative analysis method for the quality control components in Tenghuang jiangu capsules， and predict the possible action mechanism of the quality control components. METHODS Seven key quality control components in Tenghuang jiangu capsules were quantitatively analyzed by UHPLC-Q-Orbitrap HRMS. The “component-target” network was constructed based on network pharmacology， and the Kyoto Encyclopedia of Genes and Genomes （KEGG） pathway enrichment analysis and gene ontology （GO） function enrichment analysis were further conducted to find the key signaling pathways. RESULTS The average contents of succinic acid， hyperoside， gallic acid， kaempferol， naringin， naringenin and protocatechuic acid in 20 batches of Tenghuang jiangu capsules were 520.92， 67.67， 129.48， 4.74， 397.45， 5.66 and 376.62 μg/g， respectively. The results of network pharmacology showed that the 62 key target genes of the quality control components of the drug included AKT1， TNF， VEGFA， MMP9， PTGS2， etc. They were mainly enriched in cytokine receptor interaction， nuclear factor， tumor necrosis factor， interleukin 17， rheumatoid arthritis， Toll-like receptor and other signal pathways， involving inflammatory reaction， signal transduction， protein phosphorylation and other biological processes， kytoplasm， cell membrane and other cell components， as well as enzyme activity， energy activity and other molecular functions. CONCLUSIONS The established UHPLC- Q-Orbitrap HRMS method can be used for the quantitative analysis of the quality control components of Tenghuang jiangu capsule. Its quality control components may be mapped to inflammatory pathways related to bone diseases such as rheumatoid arthritis and Toll-like receptors through AKT1， TNF， VEGFA and other key targets， so as to play a therapeutic role.