Efficacy and advantage of immunotherapy for melanoma via intramuscular co-expression of plasmid-encoded PD-1 and CTLA-4 scFvs.

Immunotherapy, in the shape of immune checkpoint inhibitors (ICIs), has completely changed the treatment of cancer. However, the increasing expense of treatment and the frequency of immune-related side effects, which are frequently associated with combination antibody therapies and Fc fragment of antibody, have limited the patient's ability to benefit from these treatments. Herein, we presented the therapeutic effects of the plasmid-encoded PD-1 and CTLA-4 scFvs (single-chain variable fragment) for melanoma via an optimized intramuscular gene delivery system. After a single injection, the plasmid-encoded ICI scFv in mouse sera continued to be above 150 ng/mL for 3 weeks and reached peak amounts of 600 ng/mL. Intramuscular delivery of plasmid encoding PD-1 and CTLA-4 scFvs significantly changed the tumor microenvironment, delayed tumor growth, and prolonged survival in melanoma-bearing mice. Furthermore, no significant toxicity was observed, suggesting that this approach could improve the biosafety of ICIs combination therapy. Overall, the expression of ICI scFvs in vivo using intramuscular plasmid delivery could potentially develop into a reliable, affordable, and safe immunotherapy technique, expanding the range of antibody-based gene therapy systems that are available.

Osteosarcoma-targeted Cu and Ce based oxide nanoplatform for NIR II fluorescence/magnetic resonance dual-mode imaging and ros cascade amplification along with immunotherapy.

BACKGROUND: As the lethal bone tumor, osteosarcoma often frequently occurs in children and adolescents with locally destructive and high metastasis. Distinctive kinds of nanoplatform with high therapeutical effect and precise diagnosis for osteosarcoma are urgently required. Multimodal optical imaging and programmed treatment, including synergistic photothermal-chemodynamic therapy (PTT-CDT) elicits immunogenetic cell death (ICD) is a promising strategy that possesses high bio-imaging sensitivity for accurate osteosarcoma delineating as well as appreciable therapeutic efficacy with ignorable side-effects. METHODS AND RESULTS: In this study, mesoporous Cu and Ce based oxide nanoplatform with Arg-Gly-Asp (RGD) anchoring is designed and successfully constructed. After loading with indocyanine green, this nanoplatform can be utilized for precisely targeting and efficaciously ablating against osteosarcoma via PTT boosted CDT and the closely following ICD stimulation both in vitro and in vivo. Besides, it provides off-peak fluorescence bio-imaging in the second window of near-infrared region (NIR II, 1000-1700 nm) and Magnetic resonance signal, serves as the dual-mode contrast agents for osteosarcoma tissue discrimination. CONCLUSION: Tumor targeted Cu&Ce based mesoporous nanoplatform permits efficient osteosarcoma suppression and dual-mode bio-imaging that opens new possibility for effectively diagnosing and inhibiting the clinical malignant osteosarcoma.

Soft-tissue reconstruction with pedicled vertical rectus abdominis myocutaneous flap after total or high sacrectomy for giant sacral tumor.

BACKGROUND: The large soft-tissue defect after total or high sacrectomy for giant sacral tumor induces high incidence of wound complications. It remains a huge challenge to reconstruct the soft-tissue defect and achieve the preferred clinical outcome. METHODS: A total of 27 patients undergoing one-stage total or high sacrectomy for giant sacral tumors between 2016 and 2021 in a tertiary university hospital were retrospectively reviewed. Participants were divided into two groups. Thirteen patients underwent a pedicled vertical rectus abdominis myocutaneous (VRAM) flap reconstruction, whereas 14 patients underwent a conventional wound closure. Patient's clinical characteristics, surgical duration, postoperative complications, and outcomes were compared between the two groups. RESULTS: Patients in VRAM and non-VRAM groups were similar in baseline characteristics. The mean tumor size was 12.85 cm (range: 10-17 cm) in VRAM group and 11.79 cm (range: 10-14.5 cm) in non-VRAM group (P = 0.139). The most common giant sacral tumor is chordoma. Patients in VRAM group had a shorter length of drainage (9.85 vs 17.14 days), postoperative time in bed (5.54 vs 17.14 days), and total length of stay (19.46 vs 33.36 days) compared with patients in non-VRAM group. Patients in the VRAM group had less wound infection and debridement than patients in non-VRAM group (15.4% vs 57.1%, P < 0.001). CONCLUSIONS: This study demonstrates the advantages of pedicled VRAM flap reconstruction of large soft-tissue defects after high or total sacrectomy using the anterior-posterior approach. This choice of reconstruction is better than direct wound closure in terms of wound infection, length of drainage, and total length of stay.

Single-cell profiling of the microenvironment in human bone metastatic renal cell carcinoma.

Bone metastasis is of common occurrence in renal cell carcinoma with poor prognosis, but no optimal treatment approach has been established for bone metastatic renal cell carcinoma. To explore the potential therapeutic targets for bone metastatic renal cell carcinoma, we profile single cell transcriptomes of 6 primary renal cell carcinoma and 9 bone metastatic renal cell carcinoma. We also include scRNA-seq data of early-stage renal cell carcinoma, late-stage renal cell carcinoma, normal kidneys and healthy bone marrow samples in the study to better understand the bone metastasis niche. The molecular properties and dynamic changes of major cell lineages in bone metastatic environment of renal cell carcinoma are characterized. Bone metastatic renal cell carcinoma is associated with multifaceted immune deficiency together with cancer-associated fibroblasts, specifically appearance of macrophages exhibiting malignant and pro-angiogenic features. We also reveal the dominance of immune inhibitory T cells in the bone metastatic renal cell carcinoma which can be partially restored by the treatment. Trajectory analysis showes that myeloid-derived suppressor cells are progenitors of macrophages in the bone metastatic renal cell carcinoma while monocytes are their progenitors in primary tumors and healthy bone marrows. Additionally, the infiltration of immune inhibitory CD47+ T cells is observed in bone metastatic tumors, which may be a result of reduced phagocytosis by SIRPA-expressing macrophages in the bone microenvironment. Together, our results provide a systematic view of various cell types in bone metastatic renal cell carcinoma and suggest avenues for therapeutic solutions.

Prediction and related genes of cancer distant metastasis based on deep learning.

Cancer metastasis is one of the main causes of cancer progression and difficulty in treatment. Genes play a key role in the process of cancer metastasis, as they can influence tumor cell invasiveness, migration ability and fitness. At the same time, there is heterogeneity in the organs of cancer metastasis. Breast cancer, prostate cancer, etc. tend to metastasize in the bone. Previous studies have pointed out that the occurrence of metastasis is closely related to which tissue is transferred to and genes. In this paper, we identified genes associated with cancer metastasis to different tissues based on LASSO and Pearson correlation coefficients. In total, we identified 45 genes associated with bone metastases, 89 genes associated with lung metastases, and 86 genes associated with liver metastases. Through the expression of these genes, we propose a CNN-based model to predict the occurrence of metastasis. We call this method MDCNN, which introduces a modulation mechanism that allows the weights of convolution kernels to be adjusted at different positions and feature maps, thereby adaptively changing the convolution operation at different positions. Experiments have proved that MDCNN has achieved satisfactory prediction accuracy in bone metastasis, lung metastasis and liver metastasis, and is better than other 4 methods of the same kind. We performed enrichment analysis and immune infiltration analysis on bone metastasis-related genes, and found multiple pathways and GO terms related to bone metastasis, and found that the abundance of macrophages and monocytes was the highest in patients with bone metastasis.

Current state and challenges of emerging biomarkers for immunotherapy in hepatocellular carcinoma (Review).

Hepatocellular carcinoma (HCC) is the most prevalent form of primary liver cancer. According to the American Cancer Society, among patients diagnosed with advanced liver cancer, HCC has the sixth-highest incident rate, resulting in a poor prognosis. Surgery, radiofrequency ablation, transcatheter arterial chemoembolization, radiation, chemotherapy, targeted therapy and immunotherapy are the current treatment options available. Immunotherapy, which has emerged as an innovative treatment strategy over the past decade, is serving a vital role in the treatment of advanced liver cancer. Since only a small number of individuals can benefit from immunotherapy, biomarkers are required to help clinicians identify the target populations for this precision medicine. These biomarkers, such as PD-1/PD-L1, tumor mutational burden and circulating tumor DNA, can be used to investigate interactions between immune checkpoint inhibitors and tumors. The present review summarizes information on the currently available biomarkers used for immunotherapy and the challenges that are present.

Perspectives for immunotherapy of EBV-associated GLELC: A relatively "hot" tumor microenvironment.

BACKGROUND: Epstein-Barr virus (EBV)-associated gastric lymphoepithelioma-like carcinoma (EBVaGLELC) represents a small number of gastric cancer (GC), and research on tumor microenvironment (TME) and treatment strategy are still lacking. AIMS: Here, we aim to elucidate the immune features of this rare disease and further help to develop more effective treatment options. MATERIALS & METHODS: A retrospective analysis was conducted between 2019 to 2022 in West China Hospital to reveal the immunological characteristics of EBV-positive GLELC. The difference of immune cell subset and tumor vascular structure between gastric denocarcinoma (GAC) and EBVaGLELC will be pointed out. DISCUSSION: 13 patients with GELEC and 8 patients with GAC were retrospectively studied. The heterogeneity of the immune cell profile was then confirmed through multiplexed immunofluorescence staining (mIF), which revealed a higher proportion of CD3+ T cells, CD8+ T cells, and Treg cells in the EBV-associated GLELC group. Such a distinct TME may provide therapeutic advantages, and patients with this rare subtype of GC could be good candidates for immune checkpoint inhibitors (ICIs). Angiogenesis in EBV-positive GLELC may be less intense than that in gastric adenocarcinoma (GAC), a feature that might decrease their susceptibility to antiangiogenic therapy. Furthermore, we reported a 52-year-old male with advanced EBV-positive GLELC who showed a favorable response to the combined therapy with . A repeat evaluation showed sustained partial response (PR), and the progression-free survival (PFS) was more than 34 months until now. CONCLUSION: Compared with GAC, EBVaGLELC revealed higher T cell infiltration and less intense of angiogenesis. It displays relatively "hot" TME that may provide the rationality to treat with immunotherapy in EBV-related GLELC.

Implant failure and revision strategies after total spondylectomy for spinal tumors.

Background: Although there have been several risk factors reported for implant failure (IF), little consensus exists. Potential applicable measures to protect patients from IF are relatively few. This study aimed to discover new risk factors for IF and explore potential protective measures from IF after total spondylectomy for spinal tumors. Methods: A total of 145 patients undergoing total spondylectomy for thoracic and lumbar spinal tumors between 2010 and 2021 were included from three tertiary university hospitals. Patient demographic and surgical characteristics and follow-up outcomes were collected. Results: During a mean follow-up of 53.77 months (range, 12 to 149 months), 22 of 145 patients (15.17%) developed IF. Patients undergoing thoracolumbar junctional region (T12/L1) resection were more likely to develop IF compared to those undergoing surgery at other vertebral levels (HR = 21.622, 95% CI = 3.567-131.084, P = 0.001). Patients undergoing titanium mesh cage reconstruction were more likely to develop IF compared to patients undergoing expandable titanium cage reconstruction (HR = 8.315, 95% CI = 1.482-46.645, P = 0.016). Patients with bone cement augmentation around the cage were less likely to develop IF compared to those not receiving bone cement augmentation (HR = 0.015, 95% CI = 0.002-0.107, P < 0.001). Of the 22 patients with IF, 14 (63.63%) accepted personalized revision surgery. Conclusion: The use of an expandable cage and the use of bone cement augmentation around the anterior column support cage are protective measures against IF after total spondylectomy.

Modified Iliac Screw in Lumbopelvic Fixation After Sacral Tumor Resection: A Single-Center Case Series.

BACKGROUND: Traditional iliac screw, S2-alar iliac screw, and modified iliac screw are the 3 common techniques for lumbopelvic fixation. The application of the modified iliac technique in sacral spinal tumors has been rarely reported. OBJECTIVE: To report the feasibility and safety of modified iliac screws after sacral tumor resection and their preliminary clinical outcomes. METHODS: Twenty-seven patients who underwent sacral tumor resection with modified iliac screw fixation between August 2017 and August 2021 at our center were clinically and radiographically evaluated. RESULTS: A total of 59 iliac screws were inserted by freehand according to the anatomic landmarks. The mean operation time was 207 minutes (range, 140-435 minutes). The average estimated blood loss was 1396 mL (300-4200 mL). Computed tomography scans showed that 2 (3.4%) screws penetrated the iliac cortex, indicating a 96.6% implantation accuracy rate. There were no iatrogenic neurovascular or visceral structure complications observed. The mean minimal distances from the screw head to the skin were 24.9 and 25.8 mm on the left and right sides, respectively. The mean minimal distances from the screw head to the horizontal level of the posterior superior iliac spine were 7.9 and 8.3 mm on the left and right sides, respectively. Two patients (7.4%) underwent reoperation for wound infection. At the latest follow-up, no patient had complications of screw head prominence, pseudarthrosis, or instrument failure. CONCLUSION: The modified iliac screw is characterized by its minimal invasiveness and simplicity of placement. It is an ideal alternative for lumbopelvic fixation after sacral tumor resection.

Effects of Huhuang Burn Liniment on wound healing and changes in IL-10 and MMP-9 levels in patients with mixed hemorrhoids.

OBJECTIVE: To explore the effects of Huhuang Burn Liniment on wound healing and levels of interleukin-10 (IL-10) and matrix metalloproteinase-9 (MMP-9) in patients with mixed hemorrhoids. METHODS: The clinical data of 113 patients with mixed hemorrhoids admitted to Chongqing Sanxia Central Hospital were retrospectively collected. All patients underwent Milligan-Morgan hemorrhoidectomy, and were divided into two groups according to different postoperative treatments. Group A was treated with 1/5000 potassium permanganate sitz bath after surgery, while group B was treated with Huhuang Burn Liniment. The treatment efficacy, wound healing time, level of pain, exudation, edema, granulation scores, anal function index, levels of IL-10 and MMP-9, quality of life scores, and complications were compared between the two groups. RESULTS: The effective rate of group B (94.74%) was higher than that of group A (60.71%) (P < 0.05). Group B had shorter length of anorectal hyperbaric zone, higher anal canal resting pressure, anal canal diastolic pressure, and anal canal systolic maximum pressure (P < 0.05), lower scores of trauma pain, edema, exudation, and granulation (P < 0.05), higher IL-10 levels, and lower MMP-9 levels (P < 0.05). The complication rate of group B (8.77%) was lower than that in group A (23.21%) (P < 0.05). After treatment, group B had shorter wound healing time and higher quality of life score than group A (P < 0.05). CONCLUSION: The application of Huhuang Burn Liniment in patients with mixed hemorrhoids after surgery could promote wound healing and anal function, reduce trauma pain, exudation and edema, and improve quality of life.

3D-Printed Patient-Customized Artificial Vertebral Body for Spinal Reconstruction after Total En Bloc Spondylectomy of Complex Multi-Level Spinal Tumors.

Three dimensional (3D)-printing technology facilitates complex spine surgery with unique advantages in artificial vertebral body design and manufacturing. In this study, we aimed to demonstrate how a 3D-printed spinal implant is utilized in the management of multi-level spinal tumors and integrates with comprehensive oncologic treatment. Eight spinal or paraspinal tumor patients requiring spinal reconstruction after total en bloc spondylectomy were selected as candidates for 3D-printed titanium artificial vertebral body implants. All patients underwent surgery on three or more vertebral segments or complex spinal junction segments. The clinical, oncological, and surgical characteristics of patients were collected. Of the eight candidates, seven suffered from pain and/or limb disorder. Six underwent successful 3D-printed spinal implantation, while two failed due to implant mismatching and were converted to conventional reconstruction. Of the six patients undergoing 3D-printed spinal implant surgery: (i) Five had recurrent tumors; (ii) three underwent neoadjuvant therapy; (iii) the median surgery time was 414 min; (iv) the median blood loss was 2150 ml; (v) the median blood transfusion was 2000 ml; (vi) the median length of hospital stay was 9 days; (vii) four patients received adjuvant therapy after surgery; and (viii) all patients experienced no pain, moved freely, and had no local recurrence at a median of 11.5 months post-operative follow-up. Spinal reconstruction with a 3D-printed titanium artificial vertebral body allows for total en bloc resection of complex multi-level spinal tumors. Combined with neoadjuvant and adjuvant therapy, these patients had excellent postoperative outcomes, long-term normal spinal function, and associated low local recurrence probability.

Computer-Aided Design and 3D Printing of Hemipelvic Endoprosthesis for Personalized Limb-Salvage Reconstruction after Periacetabular Tumor Resection.

3D-printed hemipelvic endoprosthesis is an emerging solution for personalized limb-salvage reconstruction after periacetabular tumor resection. Further clinical studies are still required to report its surgical characteristics, outcomes, benefits and drawbacks. Sixteen consecutive patients underwent periacetabular tumor wide resection and pelvic reconstruction with a 3D-printed hemipelvic endoprosthesis from 2018 to 2021. The surgical characteristics and outcomes are described. The mean follow-up duration was 17.75 months (range, 6 to 46 months). Five patients underwent surgery for type I + II resection and reconstruction, seven for type II + III resection and reconstruction, three for type II resection and reconstruction, and one for type I + II + IV resection and reconstruction. The incidence of postoperative complication was 12.5% (2/16) for deep venous thrombosis (DVT), 12.5% (2/16) for pneumonia, and 12.5% (2/16) for would deep or superficial infection. During follow-up, two patients (12.5%) suffered hip dislocation and underwent revision surgery. CT demonstrated an obvious prosthetic porous structure-bone fusion after follow-up of at least 6 months. At the final follow-up, 12 lived with no evidence of disease while four lived with disease; no patients experienced pain; and 15 had independent ambulation, with a mean Musculoskeletal Tumor Society (MSTS) score of 85.8% (range, 26.7% to 100%). 3D-printed hemipelvic endoprosthesis facilitates wide resection of periacetabular tumor and limb-salvage reconstruction, thus resulting in good oncological and functional outcomes. The custom-made nature is able to well mimic the skeletal anatomy and microstructure and promote osseointegration. Perioperative complications and rehabilitation exercise still need to be stressed for this engineering technology-assisted major orthopedic surgery.

Freehand S2-Alar-Iliac Screw Placement Technique in Lumbosacral Spinal Tumors: A Preliminary Study.

OBJECTIVE: S2-alar-iliac (S2AI) screw technique is widely used in spinal surgery, but it is rarely seen in the field of spinal tumors. The aim of the study is to report the preliminary outcomes of the freehand S2AI screw fixation after lumbosaral tumor resection. METHODS: The records of patients with lumbosacral tumor who underwent S2AI screw fixation between November 2016 to November 2020 at our center were reviewed retrospectively. Outcome measures included operative time, blood loss, complications, accuracy of screws, screw breach, and overall survival. Mean ± standard deviation or range was used to present continuous variables. Kaplan-Meier curve was used to present postoperative survival. RESULTS: A total of 23 patients were identified in this study, including 12 males and 11 females, with an average age of 47.3 ± 14.5 (range,15-73). The mean operation time was 224.6 ± 54.1 (range, 155-370 min). The average estimated blood loss was 1560.9 ± 887.0 (600-4000 ml). A total of 46 S2AI screws were implanted by freehand technique. CT scans showed three (6.5%) screws had penetrated the iliac cortex, indicating 93.5% implantation accuracy rate. No complications of iatrogenic neurovascular or visceral structure were observed. The average follow-up time was 31.6 ± 15.3 months (range, 13-60 months). Two patients' postoperative plain radiography showed lucent zone around the screw. One patient underwent reoperation for wound delayed infection. At the latest follow-up, eight patients had tumor-free survival, 11 had survival with tumor, and four died of disease. CONCLUSION: The freehand S2AI screw technique is reproducible, safe, and reliable in the management of lumbosacral spinal tumors.

Customized Multilevel 3D Printing Implant for Reconstructing Spine Tumor: A Retrospective Case Series Study in a Single Center.

OBJECTIVE: To investigate the clinical efficacy and safety of 3D printed artificial vertebral body for patients who underwent multilevel total en bloc spondylectomy (TES) and analyze whether it could reduce the incidence of implant subsidence. METHODS: This is a retrospective study. From January 2017 to May 2018, eight consecutive cases with spine tumor undergoing multilevel TES were analyzed. All patients underwent X-ray and CT examinations to evaluate the stability of internal fixation during the postoperative follow-up. Demographic, surgical details, clinical data, and perioperative complications was collected. Visual analog scale, Frankel score, and spinal instability neoplastic score (SINS) classification were also recorded. RESULTS: There were six cases of primary spinal tumor and two cases of metastatic spinal tumor. All patients achieved remarkable pain relief and improvement in neurological function. Five patients underwent operation through the posterior approach, one patient underwent operation through the anterior approach and the remaining two patients through a combined anterior and posterior approach. At the last follow-up period, X-rays showed that the 3D printed artificial vertebral body of all cases matched well, and the fixation was reliable. Hardware failure such as loosening, sinking, breaking, and displacement wasn't observed during the follow-up period. CONCLUSION: 3D printed customized artificial vertebral body can provide satisfying good clinical and radiological outcomes for patients who have undergone multilevel TES.

Comparison of Surgical Outcomes Between Separation Surgery and Piecemeal Spondylectomy for Spinal Metastasis: A Retrospective Analysis.

Objective: This study aimed to compare the outcomes between piecemeal spondylectomy and separation surgery for patients with spinal metastasis. Summary of Background Data: Piecemeal spondylectomy and separation surgery are two widely-used treatment options for spinal metastasis. However, no studies have compared the surgical outcomes between both treatment modalities. Methods: Patients with spinal metastasis who underwent piecemeal spondylectomy or separation surgery between August 2017 and April 2020 at our spine center were recruited. Demographic, preoperative, perioperative, and follow-up data were collected and analyzed. Kaplan-Meier analysis and the log-rank test were used to analyze overall survival (OS) and progression-free survival (PFS) in patients with spinal metastasis. Results: Overall, 26 patients were treated with piecemeal spondylectomy, and 29 underwent separation surgery with postoperative stereotactic radiosurgery. Both groups showed significant postoperative improvements in neurological status. The piecemeal spondylectomy group had significantly more blood loss (1784.62 ± 833.64 vs. 1165.52 ± 307.38 ml) and required longer operative time (4.76 ± 0.93 vs. 3.73 ± 1.15 h) than the separation surgery group. No significant difference in OS was found between the groups (P = 0.064); however, patients in the separation surgery group experienced less local recurrence than those in the piecemeal spondylectomy group (P = 0.0014). Notably, significant differences were detected in the development of complications between the groups (P = 0.029). Conclusion: Separation surgery led to less blood loss and reduced complications and had shorter operation time than piecemeal spondylectomy. Although no significant differences were found in OS between the groups, separation surgery was associated with better PFS compared with piecemeal spondylectomy. These findings suggest that separation surgery has some advantages over piecemeal spondylectomy for patients with spinal metastatic disease.

Low-intensity pulsed ultrasound inhibits IL-6 in subchondral bone of temporomandibular joint osteoarthritis by suppressing the TGF-ß1/Smad3 pathway.

OBJECTIVE: This study aimed to provide further information on the exact mechanisms involved in the anti-inflammatory effect of low-intensity pulsed ultrasound (LIPUS) on rabbit temporomandibular joint osteoarthritis (TMJOA) on interleukin-6 (IL-6) production in subchondral bone, IL-6 production in IL-1ß stimulated via inhibition of the TGF-ß1/Smad3 pathway in mouse embryo osteoblast precursor (MC3T3-E1) cells. DESIGN: Bilateral joints were injected with type II collagenase to establish TMJOA models in two male and four female rabbits. The left joint was continuously stimulated by LIPUS, while the right joint was treated with the power off in this model. One male and two female rabbits were used as normal healthy controls without treatment. The histological features of subchondral bone were examined by Safranin-O/Fast staining. Immunohistochemistry was conducted to evaluate IL-6 expression. Then, cells were stimulated by LIPUS with IL-1ß. IL-6 expression and activity of the TGF-ß1/Smad3 pathway were evaluated by Enzyme-linked immunosorbent assay (ELISA), Immunofluorescence and Western blotting, respectively. Specific inhibition of the TGF-ß1/Smad3 pathway was conducted by transfecting with small interfering RNA (siRNA) of type II receptor (siTßRII). RESULTS: LIPUS significantly ameliorated the production of IL-6 in vitro and in vivo. Its inhibitory effect on the production of IL-6 induced by IL-1ß in MC3T3-E1 cells was partly reversed by siTßRII knockdown. CONCLUSIONS: LIPUS inhibited IL-6 production by suppressing the TGF-ß1/Smad3 pathway of subchondral bone in TMJOA. These data revealed the part of the pathways involved in the anti-inflammatory effect of LIPUS and provided a possible treatment strategy for TMJOA patients and other inflammatory diseases.

The α-Subunit of the Chloroplast ATP Synthase of Tomato Reinforces Resistance to Gray Mold and Broad-Spectrum Resistance in Transgenic Tobacco.

Chloroplast ATP synthase (cpATPase) is responsible for ATP production during photosynthesis. Our previous studies showed that the cpATPase CF1 α subunit (AtpA) is a key protein involved in Clonostachys rosea-induced resistance to the fungus Botrytis cinerea in tomato. Here, we show that expression of the tomato atpA gene was upregulated by B. cinerea and Clonostachys rosea. The tomato atpA gene was then isolated, and transgenic tobacco lines were obtained. Compared with untransformed plants, atpA-overexpressing tobacco showed increased resistance to B. cinerea, characterized by reduced disease incidence, defense-associated hypersensitive response-like reactions, balanced reactive oxygen species, alleviated damage to the chloroplast ultrastructure of leaf cells, elevated levels of ATP content and cpATPase activity, and enhanced expression of genes related to carbon metabolism, photosynthesis, and defense. Incremental Ca2+ efflux and steady H+ efflux were observed in transgenic tobacco after inoculation with B. cinerea. In addition, overexpression of atpA conferred enhanced tolerance to salinity and resistance to the fungus Cladosporium fulvum. Thus, AtpA is a key regulator that links signaling to cellular redox homeostasis, ATP biosynthesis, and gene expression of resistance traits to modulate immunity to pathogen infection and provides broad-spectrum resistance in plants in the process.

Overexpression of HIF-1alpha in Bone Marrow Mesenchymal Stem Cells Promote the Repair of Mandibular Condylar Osteochondral Defect in a Rabbit Model.

PURPOSE: The self-repair ability of temporomandibular joint (TMJ) cartilage is limited. Hypoxia-inducible factor-1 alpha (HIF-1alpha) may induce stem cells to promote chondrogenic repair. The purpose of this study was to systematically evaluate the effect of HIF-1alpha overexpression in bone marrow mesenchymal stem cells (BMSCs) combined with collagen scaffolds on the repair of TMJ condylar osteochondral defects in a rabbit model. METHODS: Osteochondral defects of 3-mm diameter × 2-mm depth were created at the right side of the mandibular condyle in 40 New Zealand white rabbits. The defect sites were treated with simple empty, collagen scaffolds (COL), BMSCs/COL, and HIF-1alpha overexpression BMSCs/COL groups. The histomorphologic features of condylar cartilage were monitored by gross examination, safranin O-fast green staining (Solarbio, Beijing, China), and immunohistochemical staining. The changes in subchondral bone were examined by microcomputed tomography. Immunofluorescence staining was used to trace the transplanted BMSCs in vivo. RESULTS: At 12 weeks postimplantation, histologic staining showed that the osteochondral defects in the simple empty and COL groups were mainly filled with fibrous tissue, whereas the BMSCs/COL and HIF-1alpha overexpression BMSCs/COL groups repaired the defect with fibrocartilage. Furthermore, the cartilage was better organized in the HIF-1alpha overexpression BMSCs/COL group compared with the BMSCs/COL group. Microcomputed tomography showed that osteochondral defects can cause abnormal hyperosteogeny in subchondral bone, and the transplantation of BMSCs, especially HIF-1alpha overexpression BMSCs, may alleviate osteosclerosis. Immunofluorescence staining showed that HIF-1alpha overexpression can promote the survival of transplanted BMSCs. CONCLUSIONS: The transplantation of HIF-1alpha overexpression BMSCs combined with a COL scaffold promotes cartilaginous repair of condylar cartilage and inhibits subchondral bone sclerosis in TMJ condylar osteochondral defect rabbits.

Inhibition of p53 DNA binding by a small molecule protects mice from radiation toxicity.

Transcription factors are attractive therapeutic targets that are considered non-druggable because they do not have binding sites for small drug-like ligands. We established a cell-free high-throughput screening assay to search for small molecule inhibitors of DNA binding by transcription factors. A screen was performed using p53 as a target, resulting in the identification of NSC194598 that inhibits p53 sequence-specific DNA binding in vitro (IC50 = 180 nM) and in vivo. NSC194598 selectively inhibited DNA binding by p53 and homologs p63/p73, but did not affect E2F1, TCF1, and c-Myc. Treatment of cells with NSC194598 alone paradoxically led to p53 accumulation and modest increase of transcriptional output owing to disruption of the MDM2-negative feedback loop. When p53 was stabilized and activated by irradiation or chemotherapy drug treatment, NSC194598 inhibited p53 DNA binding and induction of target genes. A single dose of NSC194598 increased the survival of mice after irradiation. The results suggest DNA binding by p53 can be targeted using small molecules to reduce acute toxicity to normal tissues by radiation and chemotherapy.

Growth hormone receptor promotes osteosarcoma cell growth and metastases.

Osteosarcoma (OS) is the primary bone malignancy in children and adolescents, with a high incidence of lung metastasis and poor prognosis. Here, we report that growth hormone receptor (GHR) is overexpressed in OS samples compared with osteofibrous dysplasia. We subsequently demonstrated that GHR knockdown inhibited colony formation, promoted cell apoptosis and decreased the number of cells at G2/M phase in 143B and U2OS cells. Furthermore, knockdown of GHR inhibited tumor growth in vivo. Together, these findings indicate that GHR modulates cell proliferation and metastasis through the phosphoinositide 3-kinase/AKT signaling pathway and may be suitable for use as a putative biomarker of OS.