BACKGROUND:Hypoxia is strongly associated with the development and progression of osteoarthritic chondrocyte injury,but the specific targets and regulatory mechanisms are unclear. OBJECTIVE:A machine learning approach was used to identify KDEL(Lys-Asp-Glu-Leu)receptor 3(KDELR3)as a characteristic gene for osteoarthritis hypoxia and immune infiltration analysis,to provide new ideas and methods for the treatment of osteoarthritis. METHODS:The osteoarthritis-related datasets were downloaded from the GEO database and the GSEA website to obtain hypoxia-related genes.The osteoarthritis datasets were batch-corrected and immune infiltration analyzed using R language,and osteoarthritis hypoxia genes were extracted for differential analysis.Differentially expressed genes were analyzed for GO function and KEGG signaling pathway.Weighted correlation network analysis(WGCNA)and machine learning were also used to screen osteoarthritis hypoxia signature genes,and in vitro cellular experiments were performed to validate expression and correlate immune infiltration analysis using the datasets and qPCR. RESULTS AND CONCLUSION:(1)8492 osteoarthritis genes were obtained by batch correction and principal component analysis,mainly strongly associated with immune cells such as Macrophages M2 and Mast cells resting;200 hypoxia genes were also obtained,resulting in 41 osteoarthritis hypoxia differentially expressed genes.(2)GO analysis involved mainly biological processes such as response to nutrient levels and glucocorticoids;cellular components such as lysosomal lumen and Golgi lumen;and molecular functions such as 14-3-3 protein binding and DNA-binding transcriptional activator activity.(3)KEGG analysis of osteoarthritis hypoxia differentially expressed genes was associated with signaling pathways such as PI3K-Akt,FoxO,and microRNAs in cancer.(4)The characteristic gene KDELR3 was obtained after using WGCNA analysis and machine learning screening.(5)The gene expression of KDELR3 was found to be higher in the test group than in the control group in the synovium(P=0.014)but lower in the meniscus(P=0.024)after validation by gene microarray.(6)In vitro chondrocyte assay showed that the expression of KDELR3 was higher in cartilage than in the control group(P=0.005),while KDELR3 was closely associated with Macrophages M0(P=0.014)and T cells follicular helper(P=0.014).Using a machine learning approach,we confirmed that KDELR3 can be used as a hypoxic signature gene for osteoarthritis and may intervene in osteoarthritis pathogenesis by improving hypoxia,expecting to provide a new direction for better treatment of osteoarthritis.

BACKGROUND:Ferroptosis is strongly associated with the occurrence and progression of osteoarthritis,but the specific characteristic genes and regulatory mechanisms are not known. OBJECTIVE:To identify osteoarthritis ferroptosis signature genes and immune infiltration analysis using the WGCNA and various machine learning methods. METHODS:The osteoarthritis dataset was downloaded from the GEO database and ferroptosis-related genes were obtained from the FerrDb website.R language was used to batch correct the osteoarthritis dataset,extract osteoarthritis ferroptosis genes and perform differential analysis,analyze differentially expressed genes for GO function and KEGG signaling pathway.WGCNA analysis and machine learning(random forest,LASSO regression,and SVM-RFE analysis)were also used to screen osteoarthritis ferroptosis signature genes.The in vitro cell experiments were performed to divide chondrocytes into normal and osteoarthritis model groups.The dataset and qPCR were used to verify expression and correlate immune infiltration analysis. RESULTS AND CONCLUSION:(1)12 548 osteoarthritis genes were obtained by batch correction and PCA analysis,while 484 ferroptosis genes were obtained,resulting in 24 differentially expressed genes of osteoarthritis ferroptosis.(2)GO analysis mainly involved biological processes such as response to oxidative stress and response to organophosphorus,cellular components such as apical and apical plasma membranes,and molecular functions such as heme binding and tetrapyrrole binding.(3)KEGG analysis exhibited that differentially expressed genes of osteoarthritis ferroptosis were related to signaling pathways such as the interleukin 17 signaling pathway and tumor necrosis factor signaling pathway.(4)After using WGCNA analysis and machine learning screening,we obtained the characteristic gene KLF2.After validation by gene microarray,we found that the gene expression of KLF2 was higher in the test group than in the control group in the meniscus(P=0.000 14).(5)In vitro chondrocyte assay showed that type Ⅱ collagen and KLF2 expression was lower in the osteoarthritis group than in the control group in chondrocytes(P<0.05),while in osteoarthritis ferroptosis,mast cells activated was closely correlated with dendritic cells(r=0.99);KLF2 was closely correlated with natural killer cells(r=-1,P=0.017)and T cells follicular helper(r=-1,P=0.017).(6)The findings indicate that using WGCNA analysis and machine learning methods confirmed that KLF2 can be a characteristic gene for osteoarthritis ferroptosis and may improve osteoarthritis ferroptosis by interfering with KLF2.

Objective: To investigate the effects of lactoprotein iron chelates on rats with iron deficiency anaemia (IDA), so as to provide insights into developing and utilizing novel iron supplements. Methods: Seventy weaning female SPF-graded rats of the SD strain were randomly divided into the control group (A), model group (B), ferrous sulfate group (C), lactoferrin group (D), lactoferrin iron chelate group (E), Casein oligopeptide iron chelate group (F) and whey protein oligopeptide iron chelate group (G), with 10 rats in each group. The rats in group A were fed with normal diet, and the others were fed with poor iron diet for IDA modeling. The corresponding interventions were given by intragastric administration once a day. The iron ion concentrations of group C, E, F and G were 2.0 mg/kg, and the protein and oligopeptide concentrations of group D, E, F and G were 2 000 mg/kg. Body weight and hemoglobin of rats were measured weekly during 21-day intervention. At the end, peripheral blood samples were collected, and blood routine, iron metabolism and liver function indicators were determined. Results: After the intervention, among blood routine indicators, the rats in group C, E, F and G showed elevated hemoglobin, red blood cell, mean corpuscular volume and hematocrit, and decreased free protoporphyrin and mean corpuscular hemoglobin concentration when compared with the rats in group B (all P<0.05); among iron metabolism indicators, the rats in group C, E and G showed elevated serum ferritin, the rats in group C, E, F and G showed elevated serum iron, the rats in group C, D, E, F and G showed decreased unsaturated iron binding capacity and total iron binding capacity when compared with the rats in group B (all P<0.05); among liver function indicators, the rats in group E and G showed decreased alanine transaminase when compared with the rats in group B (both P<0.05). Conclusions Lactoprotein alone could not completely improve IDA in rats compared with traditional iron supplement (ferrous sulfate). Lactoprotein iron chelate, especially whey protein oligopeptide iron chelate, could significantly improve IDA, iron reserve and liver function damage in rats.

Objective:To explore the effect of probiotics Lactiplantibacillus plantarum(LP) WCFS1 by gavage on acute necrotizing pancreatitis (ANP) and associated ileum injury in mice. Methods:Twenty-four healthy male mice were gavaged with broad-spectrum antibiotics for 3 weeks to establish microbiota-depleted mice, and then randomly divided into control group (CON), ANP model group (ANP), LP gavage group (LP) and LP gavage and ANP induced group (LP+ ANP) , with 6 mice in each group. Mice in LP and LP+ ANP group were treated by gavage of LP (1×10 9 CFU/ml, 0.2 ml/day per mouse) for 1 week, while CON and ANP were gavaged with sterile phosphate buffered saline for 1 week instead. The ANP model was induced by intraperitoneal injection with caerulein (100 μg/kg) for 10 times with 1-hour interval between two injections and the 10th injection with lipopolysaccharide(LPS) 5 mg/kg intraperitoneally, and the mice were sacrificed 2 h later. Levels of LP in stool and ileal mucosa were detected by real-time PCR; the pancreas and ileum were collected for pathological examination to observe the extent of tissue inflammation and to score the pathology. Serum amylase activities were determined by enzymatic kinetic chemistry; serum inflammators levels and intestinal permeability were detected by ELISA; levels of inflammators in pancreatic and ileal tissues were detected by real-time PCR; ileal tight-junction proteins (occludin, claudin-1 and ZO-1) were measured by immunofluorescence staining. Results:LP levels in the stool and ileal mucosa of mice were significantly increased after LP gavage, and the differences were statistically significant (913.30±39.12 vs 2.39±1.39, 23.11±0.50 vs 1.38±0.28, all P value <0.05). The pathological scores of pancreatic tissue of CON, LP, ANP and LP+ ANP group were (0.26±0.41), (0.17±0.26), (8.55±0.46) and (6.30±0.45); the serum amylase activities were (219.70±19.73), (217.60±11.30), (2896.24±98.32) and (1837.13±131.60)U/L, IL-1β were (0.87±0.28), (1.4±0.85), (67.41±6.45) and (36.33±5.65)pg/ml, IL-6 were (0.74±0.27), (0.16±0.16), (280586.12±39163.92) and (107912.75±31283.47)pg/ml, IL-10 were (35.52±5.27), (50.99±15.34), (2008.45±184.83) and (3070.35±403.71)pg/ml; the expression level of pancreatic IL-1β mRNA was 1.42±0.39, 0.95±25, 20.53±0.50 and 10.69±1.01, IL-6 mRNA was 1.31±0.44, 0.93±0.023, 21.97±1.71 and 11.54±1.75, IL-10 mRNA was 0.93±0.14, 0.75±0.15, 0.99±0.21 and 1.76±0.19; there was no significant difference between LP and CON group, and pancreatic pathological scores, serum amylase、IL-1β and IL-6 levels, and the expression level of pancreatic IL-1β and IL-6 mRNA were significantly decreased in LP+ ANP group compared with those in ANP group, while serum IL-10 levels and the expression level of pancreatic IL-10 mRNA were significantly increased compared with ANP group, and all the differences were statistically significant (all P values <0.05). The pathological scores of ileal tissue of CON, LP, ANP and LP+ ANP group were 0, 0, (3.17±0.41) and (1.67±0.52); the levels of serum DAO of CON, LP, ANP and LP+ ANP group were (0.03±0.03), (0.02±0.02), (0.50±0.05) and (0.49±0.06)ng/ml; LPS levels were (2.75±0.35), (3.74±0.28), (7.19±0.92) and (5.88±0.38)ng/ml; the expression level of ileal IL-1β mRNA was 1.21±0.20, 1.17±0.09, 1.81±0.25 and 1.63±0.21; IL-6 mRNA was 1.01±0.29, 2.83±0.42, 54.45±8.50 and 16.87±4.42; IL-10 mRNA was 1.12±0.41, 6.09±2.51, 11.65±1.47 and 29.86±2.93. There was no significant difference between LP and CON group, except that the ileal IL-10 mRNA expression was significantly higher than that of CON group. Ileal pathological scores, serum LPS levels and the expression level of ileal IL-6 mRNA were significantly lower in LP+ ANP group than those in ANP group, while the expression level of ileal IL-10 mRNA was significantly higher than that of ANP group; the expression of ileal tight junction proteins (ocludin, claudin-1, ZO-1) was significantly higher than those in ANP group, and all the differences were statistically significant (all P values <0.05). Conclusions:LP WCFS1 gavage could ameliorate the injury of pancreatic and ileal barrier in caerulein-induced ANP mice.

Objective To assess the comparative efficacy of different methods for difficult biliary cannulation in endoscopic retrograde cholangio-pancreatography(ERCP)through a network meta-analysis.Methods Randomized controlled trials(RCTs)that compared the efficacy of different adjunctive methods(early or late needle-knife technique,pancreatic guidewire-assisted technique,pancreatic stent-assisted technique,transpancreatic sphincterotomy,persistent standard cannulation technique)for difficult biliary cannulation with each other were identified.The success rate of biliary cannulation and the incidence of post-ERCP pancreatitis(PEP)were the outcomes of interest.Pairwise and network meta-analysis and ranking according to surface under the cumulative ranking curve(SUCRA)for all methods were performed.Results Eighteen RCTs were identified according to selection criteria,and 2 033 patients were enrolled.The use of transpancreatic sphincterotomy over persistent standard cannulation technique(RR=1.34,95%CI:1.02-1.77)and over pancreatic guidewire-assisted technique(RR=1.26,95%CI:1.00-1.60)significantly increased the success rate of biliary cannulation.Based on SUCRA ranking,transpancreatic sphincterotomy followed by early needle-knife techniques were ranked highest in terms of increasing the success rate of biliary cannulation.Only early needle-knife technique significantly decreased PEP rate when compared with persistent standard cannulation technique(RR=0.53,95%CI:0.30-0.94),whereas both early needle-knife techniques and transpancreatic sphincterotomy led to lower PEP rates as compared with pancreatic guidewire-assisted technique(RR=0.41,95%CI:0.17-0.99;RR=0.49,95%CI:0.25-0.96;respectively).Based on SUCRA ranking,early needle-knife technique followed by transpancreatic sphincterotomy were ranked highest for decreasing the PEP rate of biliary cannulation.Conclusions Transpancreatic sphincterotomy increases the success rate of difficult biliary cannulation in ERCP;early needle-knife technique and transpancreatic sphincterotomy are superior to other interventions in decreasing PEP rates and should be considered as a choice of difficult biliary cannulation.

Objective:To explore the application of mobile augmented reality (mAR) technology in the teaching of neuroanatomy, and to observe its effect on students' academic performance and cognitive load.Methods:By collecting and designing various neuroanatomy multimedia teaching resources (graphics, animations and videos), using augmented reality (AR) marker-based image recognition technology, the multimedia resources were placed at the tags in the traditional book pages to make the books interactive. And various multimedia resources were combined with traditional printed books through mobile devices. Forty students were randomized into the experimental group or the control group. The experimental group was taught with mAR multimedia materials, and the control group adopted traditional teaching methods. After a 6-hour course was completed, all students had a unified test, and the academic performance test and the PAAS(platform-as-a-service) cognitive load scale were used for data collection and analysis. The variance analyses (MANOVA and ANOVA) were used for significance testing.Results:One-way MANOVA test was used to determine the learning effect of mAR on academic performance and cognitive load. The results showed that there was a significant difference between the experimental group and the control group ( P<0.05). The univariate ANOVA test found that the experimental group students who learned neuroanatomy through mAR had better test scores than the control group students. In addition, compared with the control group students, the cognitive load of students in experimental group was significantly reduced, with statistical significance (all P<0.05). Conclusion:Through the teaching practice, we found that using mAR to learn neuroanatomy helps students improve their academic performance while reducing their cognitive load.

OBJECTIVE: To evaluate the efficacy of black cohosh extracts (BCE) in improving the low estrogen status induced by postoperative gonadotropin-releasing hormone agonist (GnRHa) in patients with endometriosis. METHODS: Randomized clinical controlled trial about the improvement of low estrogen status caused by GnRHa with the treatment of BCE in patients with endometriosis after laparoscopic surgery were retrieved from Medline (Ovid), PubMed, Cochrane Library, CNKI, CBMdisc, Wanfang and VIP databases before January 2020, and meta-analysis of included studies was performed by Revman 5.3 software. RESULTS: Seven randomized controlled trials involving 745 patients were included in this study. Meta-analysis results showed that the addition of BCE did not alter hormone levels of patients, including serum estradiol levels [ =1.24, 95% (-4.58, 7.08), >0.05] and luteinizing hormone levels [ =-0.02, 95% (-0.15, 0.11), >0.05]. BCE effectively improved the perimenopausal symptoms induced by low estrogen status:improving hectic fever and sweating [ =0.1, 95% (0.02, 0.47), < 0.01], reducing the occurrence of insomnia symptoms [ =0.23, 95% (0.13, 0.39), < 0.01], improving fatigue [ =0.09, 95% (0.04, 0.20), < 0.01], reducing the occurrence of vaginal dryness [ =0.04, 95% (0.01, 0.30), < 0.01]. BCE affected Kupperman's menopausal index (KMI) score 12 weeks after the surgery [ =-11.50, 95% (-20.09, -2.90), < 0.01] and KMI score 24 weeks after the surgery [ =-23.68, 95% (-39.66, -7.69), < 0.01]. CONCLUSIONS The limited evidence so far indicates that BCE could efficiently improve perimenopausal symptoms cause by low estrogen status of the patients recieved GnRHa treatment after surgery for endometriosis, but does not alter hormone levels of patients.
Cimicifuga ; Endometriosis ; Estrogens ; Female ; Humans ; Plant Extracts

Objective To reduce the risk of cervical spinal cord injury,the most medial point of the cervical intervertebral disc that the posterior percutaneous endoscopic sheath could reach was evaluated.And that could help to determine the indication of posterior percutaneous endoscopic cervical discectomy for cervical intervertebral disc herniation.Methods Cervical MRI images for 50 randomly selected patients,21 males and 29 females with ages from 20 to 60(average 33.5± 10.03 years),were analysed.All 50 patients underwent MRI examination at our institution between January 2014 and December 2017.As 50％ of the zygapophyseal joint was preserved,on the cross-section T2-weighted MRI images,when the sheath just touched the spinal cord,the intersection point of the medial wall of sheath and cervical spinal cord (Point L) was the most medial point of the posterior percutaneous endoscopy could get.The distance between Point L and the line through and tangent to the most lateral point of cervical disc border was the length of the line section DL.The distance between the middle sagittal line of the cervical disc and the line through and tangent to the most lateral point of cervical disc border was the length of the line section D.D1/D was the most medial distance ratio of the posterior percutaneous endoscopic cervical discectomy when 50％ of the lateral zygapophyseal joint was preserved.In the same way,D'1/D was the most medial distance ratio of the posterior percutaneous endoscopic cervical discectomy when 75％ of the lateral zygapophyseal joint was preserved.Results When 50％ of the lateral zygapophyseal joint was preserved,the upper limit of 95％ confidence intervals of the most medial distance ratio that the posterior percutaneous endoscopy could get were 78％,76％,81％,93％ in C3,4,C4,5,C5,6,C6,7 respectively.This meant that the most medial distance the posterior percutaneous endoscopy could get were the 78％,76％,81％,0.93％ of the length of the line section D in C3,4,C4,5,C5,6,C6,7 respectively.The most medial distance the posterior percutaneous endoscopy could get in C5,6 or C6,7 was longer than that in C3,4,C4,5.Conclusion When 50％ of the lateral zygapophyseal joint was preserved,the upper limit of the most medial distance ratio that the posterior percutaneous endoscopy should get were 78％,76％,81％,93％ in C3,4,C4,5,C5,6,C6,7 respectively.This meant that the most medial distance the posterior percutaneous endoscopy could get were the 78％,76％,81％,93％ of the length of line section D in C3,4,C4,5,C5,6,C6,7 respectively.If the resected disc was beyond this range,the cervical spinal cord should be in the risk of being injured.

Objective Retrospective study and report on cases of "symptomatic facet of residual bone mass" caused by percutaneous transforaminal endoscopic discectomy (PTED),to analysis of its causes and revision strategies.Methods Seven cases of "symptomatic facet of residual bone mass" after PTED were found in six medical centers from July 2015 to November 2017.Weintroduced the course of diagnosis and treatment,to analysis of the causes,clinical features and revision strategies of the rare complication.Results Seven patients came from different medical centers (2 cases in Ningbo No.6 Hospital and 1 case in each of the other medical centers).The average age of the subject is 67.29±9.64 years (range from 57-83 years).Among them there were 1 male and 6 female.PTED was performed for all cases with lumbar disc herniation or stenosis.The operative segments were 1 of L2,3,2 of L3,4,3 of L4,5,1 of L5S1.Symptoms occurred immediately after surgery in all cases except one after a week of operation and another one month later.Two cases were appeared symptom of contralateral irritation,and the rest were aggravated by the original symptoms.Two cerebrospinal fluid leakage caused by bone mass piercing the dural sac.The bone mass compressed the nerve root and caused 1 case of lower limb muscle weakness.Foraminoplasty was performed during PTED in all patients.After CT scan,5 cases of bone mass were found on the same side of operation,and 2 cases were in the contralateral side.The shortest time for revision was 2 days and the longest 3 months.After conservative treatment,the symptoms were relieved in only one case.Revision surgeries were performed for all the other 6 cases,2 with microendoscopic discectomy (MED),1 mobile microendoscopic discectomy (MMED),1 small incision operation,1 PTED and 1 with minimal invasive surgery of transforaminal lumbar intervertebral fusion (MIS-TLIF).The VAS scores of low back pain and leg pain was significantly relieved from 8.67±0.52 to 1.50±0.55.Conclusion FTED may lead to residual bone mass in lumbar foraminoplasty.The penetration of the bone mass block into the spinal canal can cause the compression symptoms of the corresponding segment.The patients showed the corresponding spinal canal stenosis and nerve root irritation symptoms.A revision operation is required to remove the oppressed bone mass to relieve the symptoms as soon as possible if the conservative treatment not effective.

Objective To investigate the clinical efficacy and complications of minimally invasive transforaminal lumbar-interbody fusion (TLIF) in the treatment of lumbar spondylolisthesis. Methods Total 142 patients with single level spondylolis-thesis who treated by TLIF from 2010.01 to 2015.06 were included in this study, with 68 cases in minimally invasive TLIF (MIS-TLIF) group and 74 cases in traditional open TLIF group. The general information (age, gender, isthmic or degenerative type, per-centage of slip degree, levels), operative time, blood loss, length of postoperative hospital stay, Visual Analogue Scale (VAS) of low-back pain and leg pain, and Oswestry Disability Index (ODI) were recorded and collected. The posterior height of the interverte-bralpace and segmental lordosis, reduction of spondylolisthesis and cross-sectional area of spinal canal were measured. Results There was no statistically significant difference between the two groups in age, gender ratio, percentage of slip degree, and sur-gicallevels distribution. Total of 66 cases in MIS-TLIF group and 71 cases in Open TLIF group finished 2 years follow up, and 25 cases in MIS-TLIF group and 31 cases in Open TLIF group finished 5 years follow up. The blood loss of the MIS-TLIF group was 164.7±51.7 ml, significantly lower than the open TLIF group of 239±69.3 ml(P<0.001). The length of postoperative hospital stay was 5.9 ± 1.5 days in MIS-TLIF group, significantly shorter than the open TLIF group of 7.3 ± 3.1 days(P<0.001). The operative time of MIS-TLIF and Open TLIF was 146.3±21.9 mins, 152.0±20.4 mins, respectively, and no significant differ-ence was found between them. The VAS ofback pain, leg pain, ODI in MIS-TLIF group was 1.76±1.16, 1.91±1.36 and 23.5± 7.3 at 2 years follow up, and in Open TLIF was 1.73±1.10, 1.83±1.36 and 23.8±6.7, respectively, all of them were significant-ly different to pre-operation, however, no significant difference was found between two groups. The VAS of back pain, leg pain, ODI in MIS-TLIF group was 1.73±1.21, 1.93±1.48, and 25.4±6.8 at 5years follow up, and in Open TLIF was 1.85±1.02, 1.85± 1.33 and 26.1 ± 6.5, respectively, no significant difference between twogroups. The posterior height of the intervertebral space and segmental lordosis of MIS-TLIF was 9.52±1.67 mm and 12.11°±3.44° at 2 years follow up, while the open TLIF was 9.88± 1.54 mm and 12.98 ± 3.83° , all of them were significantly different to pre-operation,however, no significant difference between two groups. The posterior height of the intervertebral space and segmental lordosis of MIS-TLIF was 9.37 ± 1.46 mm and 11.55° ± 2.77° , while the open TLIF was 9.66 ± 1.68 mm and 12.59° ± 4.23° , no significant difference between two groups. The percentage of slip degree was reduced to 5.2％±4.6％ in MIS-TLIF and 5.6％±4.3％ in open TLIF, the cross-sectional area of spinal canal was enlarged to 139.7±19.5 mm2 and 141.7±20.7 mm2, no significant difference between two groups either. Con-clusion MIS-TLIF has less blood loss, shorter postoperative hospital stay than open TLIF, and similar clinical pain and function-al outcomes. MIS-TLIF is suggested to be a safe and effective choice in the treatment of lower grade lumbar spondylolisthesis (Grade II or less).