RORγt agonist synergizes with CTLA-4 antibody to inhibit tumor growth through inhibition of Treg cells via TGF-ß signaling in cancer.

To date, the overall response rate to checkpoint blockade remains unsatisfactory, partially due to the limited understanding of the tumor immune microenvironment. The retinoic acid-related orphan receptor γt (RORγt) is the key transcription factor of T helper cell 17 (Th17) cells and plays an essential role in tumor immunity. In this study, we used JG-1, a potent and selective small-molecule RORγt agonist to evaluate the therapeutic potential and mechanism of action of targeting RORγt in tumor immunity. JG-1 promotes Th17 cells differentiation and inhibition of regulatory T (Treg) cells differentiation. JG-1 demonstrates robust tumor growth inhibition in multiple syngeneic models and shows a synergic effect with the Cytotoxic T-Lymphocyte Antigen 4 (CTLA-4) antibody. In tumors, JG-1 not only promotes Th17 cells differentiation and increases C-C Motif Chemokine Receptor 6 (CCR6)- Chemokine (C-C motif) ligand 20 (CCL20) expression, but also inhibits both the expression of transforming growth factor-ß1 (TGF-ß1) and the differentiation and infiltration of Treg cells. In summary, JG-1 is a lead compound showing a potent activity in vitro and robust tumor growth inhibition in vivo with synergetic effects with anti-CTLA-4.

Finite element analysis of the kinematic coupling effect of the joints around talus when Ponseti manipulation.

BACKGROUND: Little information was obtained from the published papers about the kinematic coupling effect between tarsal bones during Ponseti manipulation. The aim was to explore the kinematic coupling effect of the joints around talus, to investigate the kinematic rhythm and coupling relationship of tarsal joints; to clarify the pulling effect on medial ligament of the ankle during the process of Ponseti manipulation. METHODS: The model of foot and ankle was reconstructed from the Chinese digital human girl No.1 (CDH-G1) image database. Finite element analysis was applied to explore the kinematic coupling effect of the joints around talus. The distal tibia and fibula bone and the head of talus were fixed in all six degrees of freedom; outward pressure was added to the first metatarsal head to simulate the Ponseti manipulation. Kinematic coupling of each tarsal joint was investigated using the method of whole model splitting, and medial ligament pulling of the ankle was studied by designing the model of medial ligament deletion during the Ponseti manipulation. RESULTS: All the tarsal joints produced significant displacement in kinematic coupling effect, and the talus itself produced great displacement in the joint of ankle. Quantitative analysis revealed that the maximum displacement was found in the joints of talonavicular (12.01mm), cuneonavicular (10.50mm), calcaneocuboid (7.97mm), and subtalar(6.99mm).The kinematic coupling rhythm between talus and navicular, talus and calcaneus, calcaneus and cuboid, navicular and cuneiform 1 were 1:12, 1:7, 1:2 and 1:1.6. The results of ligaments pulling showed that the maximum displacement was presented in the ligaments of tibionavicular (mean 27.99mm), talonavicular (21.03mm), and calcaneonavicular (19.18 mm). CONCLUSIONS: All the tarsal joints around talus were involved in the process of Ponseti manipulation, and the strongest kinematic coupling effect was found in the joints of talonavicular, subtalar, calcaneocuboid, and cuneonavicular. The ligaments of tibionavicular, talonavicular, and calcaneonavicular were stretched greatly. It was suggested that the method of Ponseti management was a complex deformity correction processes involved all the tarsal joints. The present study contributed to better understanding the principle of Ponseti manipulation and the pathoanatomy of clubfoot. Also, the importance of cuneonavicular joint should be stressed in clinical practice.

Dieback and Leaf Spot in Box Elder (Acer negundo) Caused by Exserohilum rostratum.

Leaf spot and dieback were observed on box elder (Acer negundo) grown in a nursery in Tai'an city, Shandong Province, China, in 2019, with a disease incidence of 86%. The incidences of Exserohilum rostratum isolation were 75% from the shoots and 66.6% from the leaves of field-infected plants. Isolates were identified at the species level on the basis of morphological characteristics and through phylogenetic analysis of concatenated partial sequences of the internal transcribed spacer (ITS) region and cam, gapdh, tef1, rpb2, tub2, and his genes from the Exserohilum isolates. The effects of temperature on the mycelial growth of the Exserohilum rostratum isolates were also characterized. In greenhouse tests, seedlings inoculated with the pathogen exhibited systemic symptoms similar to those observed in the field. In pathogenicity experiments on shoots, wounded seedlings were observed to be blighted, suggesting that leaf spot and dieback may develop into more severe blight or dieback when high winds, sudden temperature decreases, or insect infestations occur. To our knowledge, this is the first report of dieback and leaf spot caused by E. rostratum on a species of A. negundo.

[Distribution and Drug Resistance of Pathogens in Oral Mucositis Associated with Chemotherapy in Patients with Malignant Hematopathy].

OBJECTIVE: To analyze the distribution and drug resistance of pathogens in oral mucositis associated with chemotherapy in hospitalized patients with malignant hematopathy, so as to provide scientific evidences for rational selection of antibiotics and infection prevention and control. METHODS: From July 2020 to June 2022, 167 patients with malignant hematopathy were treated with chemical drugs in the Department of Hematology, Hainan Hospital, and secretions from oral mucosal infected wounds were collected. VITEK2 COMPECT automatic microbial identification system (BioMerieux, France) and bacterial susceptibility card (BioMerieux) were used for bacterial identification and drug susceptibility tests. RESULTS: A total of 352 strains of pathogens were isolated from 167 patients, among which 220 strains of Gram-positive bacteria, 118 strains of Gram-negative bacteria and 14 strains of fungi, accounted for 62.50%, 33.52% and 3.98%, respectively. The Gram-positive bacteria was mainly Staphylococcus and Streptococcus, while Gram-negative bacteria was mainly Klebsiella and Proteus. The resistance of main Gram-positive bacteria to vancomycin, ciprofloxacin and gentamicin was low, and the resistance to penicillin, cefuroxime, ampicillin, cefotaxime, erythromycin and levofloxacin was high. The main Gram-negative bacteria had low resistance to gentamicin, imipenem and penicillin, but high resistance to levofloxacin, cefotaxime, cefuroxime, ampicillin and vancomycin. The clinical data of oral mucositis patients with oral ulcer (severe) and without oral ulcer (mild) were compared, and it was found that there were statistically significant differences in poor oral hygiene, diabetes, sleep duration less than 8 hours per night between two groups (P<0.05). CONCLUSION: Gram-positive bacteria is the main pathogen of oral mucositis in patients with malignant hematopathy after chemotherapy. It is sensitive to glycopeptide antibiotics and aminoglycosides antibiotics. Poor oral hygiene, diabetes and sleep duration less than 8 hours per night are risk factors for oral mucositis with oral ulcer (severe).

Intrathecal injection of metabotropic glutamate receptor subtype 3 and 5 agonist/antagonist attenuates bone cancer pain by inhibition of spinal astrocyte activation in a mouse model.

BACKGROUND: Astrocytes and metabotropic glutamate receptors play important roles in nociceptive processing. However, their roles in bone cancer pain were not well understood. This study sought to investigate whether selective mGluR3 and mGluR5 agonist or antagonist develop antinociceptive effects on bone cancer pain by inhibition of spinal astrocyte activation. METHODS: C3H/HeNCrlVr mice were inoculated into the intramedullary space of the femur with sarcoma NCTC 2472 cells to induce bone cancer pain. Quantitative real-time reverse transcription-polymerase chain reaction and Western blot experiments examined messenger RNA and protein expression of spinal glial fibrillary acidic protein, mGluR3, and mGluR5. The authors further investigated effects of intrathecal treatment with the mGluR3 agonist (APDC), the mGluR3 antagonist (LY341495), the mGluR5 agonist (CHPG), or the mGluR5 antagonist (MTEP) on nociceptive behaviors and spinal astrocyte activation associated with bone cancer pain. RESULTS: Inoculation of sarcoma cells, but not α-MEM solution, induced progressive bone cancer pain and resulted in up-regulation of glial fibrillary acidic protein, mGluR3, and mGluR5 expression on days 10, 14, and 21 postinoculation. Intrathecal administration of APDC and MTEP attenuated bone cancer-evoked spontaneous pain, mechanical allodynia, thermal hyperalgesia, and reduced spinal glial fibrillary acidic protein expression. However, treatment with LY341495 and CHPG induced thermal hyperalgesia and spinal glial fibrillary acidic protein expression in control mice. CONCLUSIONS: Spinal mGluR3 activation or mGluR5 inhibition reduced bone cancer pain. Inhibition of spinal astrocyte activation may contribute to the analgesic effects. These findings may lead to novel strategies for the treatment of bone cancer pain.

[Role of cannabinoid 2 receptor in the development of bone cancer pain].

OBJECTIVE: To explore the effects of cannabinoid 2 receptor (CB2) in the development of bone cancer pain in mice. METHODS: A total of 84 mice (C3H/HeJ) were randomly divided into 4 groups:tumor group (Group T, n = 30), medication administration group (Group J, n = 12), vehicle group (Group D, n = 12) and sham group (Group S, n = 30). And 2 × 10(5) osteolytic NCTC2472 cells in α-MEM were injected into medullary cavity of right distal femur to induce bone cancer pain in a murine model while sham mice received an injection of only α-MEM. All mice were tested for pain-related behaviors at pre-inoculation and at Days 5, 7, 10 and 14 post-inoculation. The tests included paw withdrawal mechanical threshold (PWMT) and paw withdrawal thermal latency (PWTL). Group J and Group D were injected intrathecally with 2 µg JWH015 dissolved in 4% DMSO and only 4% DMSO respectively in a 5 µl. volume. Pain behavior tests were performed before and at 1, 6, 24, 48 and 72 h after an intrathecal injection. Lumbar intumescentia of mice in each group were harvested to examine the expression level of CB2 by Western blot after pain behavior tests at Days 5, 7, 10 and 14 post-inoculation and 12 h after an intrathecal injection. RESULTS: (1) Pain behavior tests:Mechanical allodynia appeared at Day 7 post-inoculation. The value of PWMT was (1.27 ± 0.28) g (P < 0.05) and it declined gradually to (0.53 ± 0.20) g at Day 14. The threshold of mechanical hyperalgesia increased to (1.00 ± 0.20) g at 6 h after an intrathecal injection of JWH015, peaked at (1.40 ± 0.39) g at 12 h, became alleviated after 48 h and recovered to the pre-dosing levels at 72 h. Thermal hyperalgesia appeared at Day 10 post-inoculation. The value of PWTL was (16.9 ± 0.4) s (P < 0.05) at Day 10 and declined to (11.5 ± 0.7) s at Day 14 post-inoculation. The threshold of thermal hyperalgesia increased to (15.7 ± 1.9) g at 6 h after an intrathecal injection of JWH015, peaked at (18.6 ± 2.3) g at 12 h, became alleviated after 48 h and recovered to the pre-dosing levels at 72 h. (2) Western blot: From Day 5 post-inoculation, the ratio of CB2/ß-actin increased gradually. Compared with the ratio of 0.190 ± 0.010 at Day 5 post-inoculation, the ratio of CB2/ß-actin increased to 0.660 ± 0.010 at Day 14 post-inoculation (P < 0.05); compared with the ratio of 0.903 ± 0.006 in group D at 12 h after an intrathecal injection of JWH015, the ratio of CB2/ß-actin 0.510 ± 0.010 significantly decreased (P < 0.05). CONCLUSION: The cannabinoid 2 receptor plays an important role in the formation of bone cancer pain.

Effect of Achilles tendon on kinematic coupling relationship between tarsal bones: a pilot finite element study.

BACKGROUND: The procedure of percutaneous Achilles tenotomy (PAT) is an important component of the Ponseti method. However, few studies reported the influence of Achilles tendon on kinematic coupling relationship between tarsal bones. The purpose of present study was to demonstrate the effect of Achilles tendon on the kinematic coupling relationship between tarsal bones, and to illustrate how kinematic coupling relationship between tarsal bones works in term of finite element analysis. METHODS: A three-dimensional finite element model of foot and ankle was constructed based on the Chinese digital human girl No.1 (CDH-G1) image database using the software of mimics, Geomagic studio, HyperMesh, and Abaqus. The last manipulation of the Ponseti method before the procedure of PAT was simulated. The talus head and the proximal tibia and fibula bone were fixed in all six degrees of freedom, and the outward pressure was added on the first metatarsal head to investigate the kinematic coupling relationship between tarsal bones. RESULTS: The least relationship of kinematic coupling between tarsal bones was found in calcaneus. Stress concentration was mainly observed at the navicular, talus and the medial malleolus. The difference in displacement of the navicular was only found with the Achilles tendon stiffness of 0 N/mm and others. No difference in the navicular displacement was found in the stiffness of Achilles tendon between 40, 80, 200, 400, and 1000 N/mm. The maximum displacement of navicular was observed at the ankle position of PF-20° (plantar flexion-20°). The difference in displacement of the navicular was greater at the ankle position of PF-20° with the Achilles tendon stiffness of 0 N/mm than that at the ankle position of PF-40° with the Achilles tendon stiffness of 40 N/mm. CONCLUSIONS: Based on the findings from this study, it was demonstrated that the Achilles tendon existence or not and ankle position had great influence, while increased stiffness of Achilles tendon had no influence on kinematic coupling relationship between tarsal bones. For the cases with severe equinus, earlier implementation of PAT procedure (with the purpose of release the Achilles tendon and reduce the degree of ankle plantar flexion) may be beneficial to the deformity correction.

Long non­coding RNA BC168687 small interfering RNA reduces high glucose and high free fatty acid­induced expression of P2X7 receptors in satellite glial cells.

Purinergic signaling contributes to inflammatory and immune responses. The activation of the P2X purinoceptor 7 (P2X7) in satellite glial cells (SGCs) may be an essential component in the promotion of inflammation and neuropathic pain. Long non­coding RNAs (lncRNAs) are involved in multiple physiological and pathological processes. The aim of the present study was to investigate the effects of a small interfering RNA for the lncRNA BC168687 on SGC P2X7 expression in a high glucose and high free fatty acids (HGHF) environment. It was demonstrated that BC168687 small interfering (si)RNA downregulated the co­expression of the P2X7 and glial fibrillary acidic protein and P2X7 mRNA expression. Additionally, HGHF may activate the mitogen­activated protein kinase signaling pathway by increasing the release of nitric oxide and reactive oxygen species in SGCs. Taken together, these results indicate that silencing BC168687 expression may downregulate the increased expression of P2X7 receptors in SGCs induced by a HGHF environment.

Detection of gene expression pattern in the early stage after spinal cord injury by gene chip.

OBJECTIVE: To study the changes of the gene expression pattern of spinal cord tissues in the early stage after injury by DNA microarray (gene chip). METHODS: The contusion model of rat spinal cord was established according to Allen's falling strike method and the gene expression patterns of normal and injured spinal cord tissues were studied by gene chip. RESULTS: The expression of 45 genes was significantly changed in the early stage after spinal cord injury, in which 22 genes up-regulated and 23 genes down-regulated. CONCLUSIONS: The expression of some genes changes significantly in the early stage after spinal cord injury, which indicates the complexity of secondary spinal cord injury.

[Surgical therapy of lumbar disk herniation in 6 815 cases].

OBJECTIVE: To explore the surgical therapy for lumbar disc herniation (LDH). METHODS: Under local anesthesia by blocking the posterior branches of spinal nerves, 3 surgical approaches were adopted in 6 815 LDH cases, namely discectomy by lateral and interlamina access, double fenestration and enhanced decompression for broad central LDH, and removal of nucleus pulposus with small incision and fenestration. RESULTS: Follow-up study was conducted in 5 324 cases for the duration varying from 6 months to 14 years, averaging 6.3 years. Excellent results were achieved in 4 260 cases (80.02%), good in 855 cases (16.06%) with another 209 cases (3.92%) showing symptomatic improvement. X-ray examination in these cases revealed narrowed intervertebral spaces of different degrees without lumbar instability or spondylolisthesis. CONCLUSION: In comparison with conventional approaches, the 3 surgical approaches for LDH management may preserve the utmost integrity of the posterior lumbar structures and cause less traumatic injuries with shortened operation time and quick recovery.

[Clinical treatment of diabetes mellitus complicated with chronic osteomyelitis].

OBJECTIVE: To investigate treatment methods for diabetes complicated with chronic osteomyelitis. METHODS: The clinical data of 39 diabetic patients with chronic osteomyelitis from 2000 to 2011 were studied. Among the patients, 23 patients were male and 16 patients were female,ranging in age from 21 to 81 years old,with a mean of 45 years old. All the patients had symptoms such as red,swelling,hot and pain to different degrees,and the duration of the disease lasted over more than 3 months,among which,25 patients had preoperative sinus formation. Routine preoperative drainage fluid culture, X-ray and CT examination were performed. Twenty-seven patients underwent debridement, one stage bone grafting and external fixation; 12 patients underwent debridement and antibiotic treatment. The examination results of ESR, CRP and blood routine before and after treatment were compared to evaluate clinical effects. RESULTS: After treatment, the values of ESR(preoperative 35 patients with higher value, and postoperative 32 patients with normal value), blood routine (preoperative 25 patients with abnormal value,and postoperative 23 patients with normal value), and CRP(preoperative 36 patients with higher value, and postoperative 33 patients with normal value) all improved. The liquor puris cultures were negative after treatment. Among 39 patients, 33 patients had clinical cure; 6 patients recurred within 4 months to 4 years. CONCLUSION: The treatment of diabetes should be paid attention when the chronic osteomyelitis was treated,and the combination treatment can significantly improve the cure rate of diabetes complicated with chronic osteomyelitis.

[Investigation of articular cavity medicating for treatment of knee osteoarthritis].

Osteoarthritis (OA) is a common chronic progressing disease in the elderly. In current there are many methods of drug treatment for knee osteoarthritis, the intra-articular injection belongs to local application, can direct effect on arthritis, is the most used and most effective methods in osteoarthritis treatment. A single medication may exist that anti-inflammatory effects produce a marked effect but articular cartilage injury is induced, or articular cartilage repairing are promoted but anti-inflammatory effects is not ideal. Therefore,the best method of treatment should be a comprehensive analysis of articular structure, function and possible pathogenic factor to understand the expectations of level of recovery and use in patients to design an individualized treatment plan.

Association of genetic variants in the IRAK-4 gene with susceptibility to severe sepsis.

BACKGROUND: The association of genetic variation in the IRAK-1 gene with sepsis outcome has been proved. However, few studies have addressed the impact of the IRAK-4 gene variants on sepsis risk. This study aimed to determine whether the polymorphisms in the IRAK-4 gene are associated with susceptibility to and prognosis of severe sepsis in the Chinese Han ethnic population. METHODS: In this case-control study, 192 patients with severe sepsis hospitalized in the emergency department of Zhongshan Hospital from February 2006 to December 2009 and 192 healthy volunteers were enrolled. Exclusion criteria included metastatic tumors, autoimmune diseases, AIDS or treatment with immunosuppressive drugs. This study was approved by the ethical committee of Zhongshan Hospital, Fudan University. Sepsis patients were divided into a survival group (n=124) and a non-survival group (n=68) according to the 30-day mortality. Primer 3 software was used to design PCR and sequencing primers. Genomic DNA was extracted from peripheral blood mononuclear cells. Seven tagSNPs in IRAK-4 were selected according to the data of the Chinese Han population in Beijing from the Hapmap project and genotyped by direct sequencing. The chi-square test was used to evaluate the differences in genotype and allele frequencies between the two groups. RESULTS: The distributions of all tagSNPs were consistent with Hardy-Weinberg equilibrium. The allele and genotype frequencies of rs4251545 (G/A) were significantly different between the severe sepsis and healthy control groups (P=0.015, P=0.035, respectively). Carriers of the rs4251545A had a higher risk for severe sepsis compared with carriers of the rs4251545G (OR=1.69, 95% CI: 1.10-2.58). The allele and genotype frequencies of all SNPs were not significantly different between the survival group and non-survival group. CONCLUSION: These findings indicate that the variants in IRAK-4 are significantly associated with susceptibility to severe sepsis in the Chinese Han ethnic population.

[Pullout test in expansive unilateral open-door laminoplasty of the cervical spine with OsteoMed M3 plate and screws: the screw orientation matters].

OBJECTIVE: To study the impact of screw orientation on the pullout strength of OsteoMed M3 titanium screws in expansive unilateral open-door laminoplasty of the cervical spine. METHODS: Six fresh human cervical spine specimens were randomly numbered and OsteoMed M3 plate and screws were used for an expansive unilateral open-door laminoplasty. The screws were inserted in the lateral mass at different extraversion angles (0°, 30° and 45°). The maximum pullout strength was tested on the ElectroForce material testing machine. RESULTS: The maximum pullout strength was 81.60∓7.33 N, 150.05∓15.57 N, and 160.08∓17.77 N in extraversion angle 0°, 30°, and 45° groups, respectively. The maximum pullout strength was significantly less in extraversion angle 0° group than in 30° and 45° groups (P<0.05), but similar in the latter two groups. CONCLUSION: The pullout strength of the screws inserted at an extraversion angle over 30° provides stronger fixation than an angle of 0° in the unilateral open-door laminoplasty using OsteoMed M3 titanium plate and screws.

[Expression of hypoxia-inducible factor-1α and vascular endothelial growth factor in the synovium of patients with osteoarthritis].

OBJECTIVE: To detect the expression of hypoxia-inducible factor-1α (HIF-1α) and vascular endothelial growth factor (VEGF) in patients with osteoarthritis and investigate their roles in the synovial lesions of osteoarthritis. METHODS: The expressions of HIF-1α and VEGF in the synovium were detected by immunohistochemical staining in 30 osteoarthritis cases, 20 acute traumatic arthritis cases and 10 normal synovial biopsy samples. The correlation between HIF-1α and VEGF, and their relationships with osteoarthritis were analyzed. RESULTS: The rates of positive expression of HIF-1α and VEGF in osteoarthritis cases were significantly higher than those in acute traumatic arthritis (86.7% vs 60% and 80% vs 48%, P<0.05). Normal human synovium showed no positive expressions of HIF-1α and VEGF. HIF-1α expression was positively correlated to VEGF expression in acute traumatic synovitis and osteoarthritis cases, with correlation coefficients of 0.666 and 0.678, respectively. CONCLUSION: The expressions of HIF-1α and VEGF in the synovial tissue are significantly higher in osteoarthritis cases than in cases of acute traumatic arthritis. They have close relationship in the synovial lesions of osteoarthritis and both contribute to the development of osteoarthritis.

[Interface shear stress between the artificial bones and injectable calcium phosphate glue: an experimental study in rabbits].

OBJECTIVE: To evaluate the effect of the composite bone material, fibrin glue (FG) combined with beta-tricalcium phosphate (beta-TCP)/monocalcium phosphate, in repairing bone defects and assess the feasibility of using this cement for artificial joint fixation. METHODS: Bone defects were induced in 16 normal adult New Zealand white rabbits at the bilateral femoral lateral condyles where an 8-mm-deep hole (4 mm in diameter) was drilled on each side. The composite FG/calcium phosphate cement (CPC) (solution: power volume ratio of 0.3:1) was injected on one side of the bone defects (experimental group) and pressurized for 10 minutes, and CPC was injected on the other side (control). The rabbits were sacrificed at 2, 4, 8 or 12 weeks after the operation for gross observation and biomechanical tests. RESULT: The composite material FG/CPC was more effective than CPC for bone defect repair, and biomechanical tests revealed significant differences between them (P<0.05). The shear stress of the artificial bone cement in the FG/CPC group was stronger than that in CPC group. CONCLUSION: The FG/CPC composite possesses good biocompatibility and osteoconductivity and may serve as an ideal material for repairing bone defects.