BdCV1-Encoded P3 Silencing Suppressor Identification and Its Roles in Botryosphaeria dothidea, Causing Pear Ring Rot Disease.

Pear ring rot disease is an important branch disease, caused by Botryosphaeria dothidea. With the discovery of fungal viruses, the use of their attenuated properties for biological control provides a new strategy for the biological control of fungal disease. RNA silencing is a major antiviral defense mechanism in plants, insects, and fungi. Viruses encode and utilize RNA silencing suppressors to suppress host defenses. Previous studies revealed that Botryosphaeria dothidea chrysovirus 1 (BdCV1) exhibited weak pathogenicity and could activate host gene silencing by infecting B. dothidea. The aim of our study was to investigate whether BdCV1 can encode a silencing suppressor and what effect it has on the host. In this study, the capability of silencing inhibitory activity of four BdCV1-encoded proteins was analyzed, and the P3 protein was identified as a BdCV1 RNA silencing suppressor in the exotic host Nicotiana benthamiana line 16c. In addition, we demonstrated that P3 could inhibit local silencing, block systemic RNA silencing, and induce the necrosis reaction of tobacco leaves. Furthermore, overexpression of P3 could slow down the growth rate and reduce the pathogenicity of B. dothidea, and to some extent affect the expression level of RNA silencing components and virus-derived siRNAs (vsiRNAs). Combined with transcriptomic analysis, P3 had an effect on the gene expression and biological process of B. dothidea. The obtained results provide new theoretical information for further study of interaction between BdCV1 P3 as a potential silencing suppressor and B. dothidea.

The role of circadian clock genes in colorectal carcinoma: Novel insights into regulatory mechanism and implications in clinical therapy.

Colorectal cancer (CRC) is a lethal malignancy with limited treatment strategies. Accumulating evidence indicates that CRC tumorigenesis, progression and metastasis are intimately associated with circadian clock, an inherent 24-h cycle oscillation of biochemical, physiological functions in almost every eukaryote. In the present review, we summarize the altered expression level of circadian genes in CRC and the prognosis associated with gene abundance switch. We illustrate the function and potential mechanisms of circadian genes in CRC pathogenesis and progression. Moreover, circadian based-therapeutic strategies including chronotherapy, therapeutics targeting potential circadian components, and melatonin treatment in CRC are also highlighted.

The PcMYB44-mediated miR397-PcLACs module regulates defence-induced lignification in pear resistance to fungal disease.

Diseases caused by Alternaria alternata and Botryosphaeria dothidea diminish pear yield and quality, and restrict the pear agricultural industry. Lignification is a conserved mechanism for plant resistance against pathogen invasion. The regulatory mechanisms underlying defence-induced lignification in pear in response to fungal pathogen infection remain unknown. In this study, analysis of lignification level and lignin content in pear revealed that A. alternata and B. dothidea induced lignification, and transcriptomics showed that lignin biosynthesis was affected. To explore whether laccases (LACs) mediated by miR397 regulate lignification in pear, we investigated the role of PcmiR397 in repressing the expression of PcLACs using 5'-RNA ligase-mediated-RACE and co-transformation in tobacco. Opposite expression patterns for PcmiR397 and PcLAC target genes were observed in pear in response to pathogens. Transient transformation in pear demonstrated that silencing PcmiR397 and overexpressing a single PcLAC enhanced resistance to pathogens via lignin synthesis. To further reveal the mechanism underpinning the PcMIR397 response of pear to pathogens, the PcMIR397 promoter was analysed, and pMIR397-1039 was found to be inhibited by pathogen infection. The transcription factor PcMYB44 was up-regulated, and it bound to the PcMIR397 promoter and inhibited transcription following pathogen infection. The results demonstrate the role of PcmiR397-PcLACs in broad-spectrum resistance to fungal disease, and the potential role of PcMYB44 involved in the miR397-PcLAC module in regulating defence-induced lignification. The findings provide valuable candidate gene resources and guidance for molecular breeding to improve resistance to fungal disease in pear.

Colorectal cancer: Metabolic interactions reshape the tumor microenvironment.

Colorectal cancer (CRC) is one of the most common cancers worldwide, which ranks third in terms of incidence and the second leading cause of cancer-related mortality. Metabolic reprogramming within the tumor microenvironment (TME) has been proved intimately involved in the initiation and malignant progression of CRC. Signal messengers, including cytokines, metabolites, and exosomes among others, derived from cancer cells can be utilized by the surrounding cells within the TME to induce metabolic alteration and cancer-associated transformation. In turn, the cargos secreted from cancer-associate cells further provide the nutrition and energy supply for cancer cells, supporting their metabolic reprogramming to promote proliferation, migration, metastasis, and radiochemoresistance. In this review, we focus on the main cellular components in the TME: CAFs, TAMs, lymphocytes and neutrophils, and enumerate and integrate how the metabolic interactions between these components and cancer cells reshape TME to foster CRC malignancy.

Molecular characterization of a new fusarivirus infecting Botryosphaeria dothidea, the causal agent of pear ring rot disease.

Here, a novel mycovirus, tentatively designated as "Botryosphaeria dothidea fusarivirus 2" (BdFV2), was discovered in Botryosphaeria dothidea strain JZ-3. The complete genome sequence is 6,271 nucleotides (nt) in length, excluding the poly(A) tail, and contains two putative open reading frames (ORFs). The larger ORF1 encodes a polypeptide of 1,552 amino acids (aa) with conserved RNA-dependent RNA polymerase (RdRp) domains and a viral helicase domain. The ORF1-encoded polypeptide shares 19.47-78.70% sequence identity with those of other fusariviruses and shares the highest sequence identity (78.70%) with the corresponding protein aa sequences of Neofusicoccum luteum fusarivirus 1 (NlFV1) isolate CBS110299. The small ORF2 encodes a hypothetical protein with 479 aa, which is predicted to contain a chromosome segregation protein SMC domain of unknown function. Sequence alignments and phylogenetic analysis indicated that BdFV2 is a distinct member of the recently established family Fusariviridae. BdFV2 appears to be a novel fusarivirus infecting a pathogenic B. dothidea strain that causes pear ring rot disease.

Preparation and formation mechanism of biomass-based graphite carbon catalyzed by iron nitrate under a low-temperature condition.

Graphite is a widely used industrial material, which experienced a marked shortage caused by the growing demand for electrode anode material and the increased costs for raw material. Graphitic carbon from biomass is a promising approach that will result in low-cost and efficient preparation. Herein, Fe(NO3)3 was selected as the catalyst for pine sawdust, and the effects of temperature and iron content on the graphitization of biochar were investigated. Additionally, the formation mechanism of the graphitic crystallite structure was explored. Results showed that the formation of pyrolysis gas increased with the increase in the amount of catalyst added or pyrolysis temperature. The change in pyrolysis gas, such as H2 and CO, was a critical auxiliary factor reflecting the conversion process. As temperature was increased from 600 °C to 800 °C, the solid products showed high graphitization and low solid yield. Graphite structure mainly formed at 700 °C because of the formation of Fe nanoparticles. The increase in the amount of catalyst could provide more reaction sites and promote the contact between Fe and C, showing that amorphous carbon is dissolved on Fe nanoparticles and precipitated into ordered graphitic carbon. On this basis, a mechanism of "carbon dissolution-precipitation" was proposed to explain the formation of graphite structure, and the whole pyrolysis process included the transformation of the iron element were analyzed.

Is frozen tumour-bearing autograft with concurrent vascularized fibula an alternative to the Capanna technique for the intercalary reconstruction after resection of osteosarcoma in the lower limb?

AIMS: The use of frozen tumour-bearing autograft combined with a vascularized fibular graft (VFG) represents a new technique for biological reconstruction of massive bone defect. We have compared the clinical outcomes between this technique and Capanna reconstruction. METHODS: From June 2011 to January 2016 a retrospective study was carried out of patients with primary osteosarcoma of lower limbs who underwent combined biological intercalary reconstruction. Patients were categorized into two groups based on the reconstructive technique: frozen tumour-bearing autograft combined with concurrent VFG (Group 1) and the Capanna method (Group 2). Demographics, operating procedures, oncological outcomes, graft union, limb function, and postoperative complications were compared. RESULTS: A total of 23 patients were identified for analysis: eight in Group 1 and 15 in Group 2. There was no difference in the demographics (age, sex, and affected site) and operating procedures (resection length, duration of surgery, and blood loss) between the two groups. No significant difference was found in local recurrence in Group 1 versus Group 2 (p = 0.585). Mean union time for the frozen autograft-host junction was 8.4 months (7.0 to 11.0), significantly earlier than for the allograft-host junction in Group 2 (mean 14.1 months (10.0 to 28.0); p < 0.001). Mean Musculoskeletal Tumor Society scores in groups 1 and 2 were 90.3% (SD 7.4%) and 88.0% (SD 9.0%), respectively, with no significant statistical difference (p = 0.535). In terms of complications, infection (n = 1, 6.7%) and delayed union (n = 2, 13.3%) occurred in Group 2, but no such complications were observed in Group 1. CONCLUSION: Frozen tumour-bearing autograft in combination with VFG can be used as an alternative to the Capanna reconstruction in properly selected patients with osteosarcoma. Cite this article: Bone Joint J 2020;102-B(5):646-652.

Factors Influencing Osseous Union Following Surgical Treatment of Bone Tumors with Use of the Capanna Technique.

BACKGROUND: The Capanna technique involves the use of a vascularized fibular graft inlaid in a massive bone graft in intercalary reconstruction for diaphyseal long-bone defects caused by tumor resection. Allograft-host union time varies in different reports, and few studies have focused on the underlying factors affecting union time. The purpose of the present study was to analyze factors relevant to union time and to report complications of the Capanna technique. METHODS: We identified 60 patients who underwent segmental reconstruction with use of the Capanna technique following tumor resection (in the humerus in 10 patients, the femur in 33 patients, and the tibia in 17 patients). Multivariable linear multiple regression model analysis was performed with allograft-host osseous union time as the dependent variable. Union time was evaluated on radiographs. Independent variables included age, tumor site, adjuvant treatment, a previous surgical procedure, defect length, fixation method, and fibular viability. A retrieved specimen of the composite was histologically assessed. RESULTS: The mean defect length was 16 cm. All allografts and host bone united, with the mean time to union of 13 months (range, 6 to 27 months). Prolonged union time was associated with devitalization of the fibular graft (p < 0.001), use of chemotherapy (p = 0.031), and a previous surgical procedure (p = 0.048). Patient age (p = 0.742), amount of resection (p = 0.907), operative site (p = 0.508), and fixation method (p = 0.105) were not associated with union time. On histological analysis, we found that the allograft-host cortical junction was united by callus from both periosteum of the host bone and the fibula. CONCLUSIONS: The Capanna technique appears to be a reliable method for intercalary reconstruction with a low rate of complications. Devitalization of the transplanted fibula, chemotherapy, and a previous surgical procedure are adverse factors leading to prolonged union time. LEVEL OF EVIDENCE: Therapeutic Level IV. See Instructions for Authors for a complete description of levels of evidence.

Gastrodia elata Blume Polysaccharides: A Review of Their Acquisition, Analysis, Modification, and Pharmacological Activities.

Gastrodia elata Blume (G. elata) is a valuable Traditional Chinese Medicine (TCM) with a wide range of clinical applications. G. elata polysaccharides, as one of the main active ingredients of G. elata, have interesting extraction, purification, qualitative analysis, quantitative analysis, derivatization, and pharmacological activity aspects, yet a review of G. elata polysaccharides has not yet been published. Based on this, this article summarizes the progress of G. elata polysaccharides in terms of the above aspects to provide a basis for their further research and development.

What are the Oncologic and Functional Outcomes After Joint Salvage Resections for Juxtaarticular Osteosarcoma About the Knee?

BACKGROUND: Joint salvage surgery for patients with juxtaarticular osteosarcoma remains challenging, especially when the tumor invades the epiphysis. Because patients are surviving longer with current chemotherapy regimens, it is advantageous to retain native joints if possible, especially in young patients. However, the results using joint-preserving tumor resections in this context have not been well characterized. QUESTIONS/PURPOSES: (1) What are the functional outcomes after limb salvage surgery at a minimum of 3 years? (2) What are the oncologic outcomes? (3) Is joint salvage surgery for epiphyseal tumors associated with an increased risk of local recurrence compared with metaphyseal tumors not invading the epiphysis? (4) What are the complications associated with joint salvage surgery? METHODS: Between 2004 and 2013, we treated 117 patients with juxtaarticular osteosarcoma; of those, 43 (38%) were treated with joint salvage surgery, and 41 (95%) of the 43 patients are included in our study. The other two (5%) were lost to followup before 3 years (mean, 4.4 years; range, 3-11 years,). During the period in question, we generally performed joint salvage surgery in these patients when they had a favorable response to chemotherapy, did not have a pathologic fracture or extrusion of the tumor into the joint, and did not have a whole-epiphyseal osteolytic lesion, a large mass, or obvious neurovascular involvement. This report is a followup of an earlier study; the current study includes an additional nine patients, and additional followup of a mean of 19 months for the patients included in the earlier report. We ascertained overall survival and survival free from local recurrence which was estimated using the Kaplan-Meier method, functional status of the limb which was evaluated using the Musculoskeletal Tumor Society (MSTS)-93 scoring system, and recorded reconstructive complications including infection, fracture, skin necrosis, and nonunion. We compared oncologic and functional outcomes between patients with (n = 28) and without tumor extension to epiphysis (n =13). We also compared oncologic and functional outcomes among patients with different adjuvant treatments including microwave ablation (n = 11), cryoablation (n = 12), and navigation-assisted osteotomy (n = 5). Complications were tallied using records from our institutional database. RESULTS: The overall Kaplan-Mayer survival rate was 82% (95% CI, 104-128 months) at 5 years. The overall Kaplan-Meier survivorship from local recurrence was 91% at 5 years (95% CI, 115-133 months). Three patients had a local recurrence, but none had local recurrence in or close to the remaining epiphysis. The MSTS scores ranged from 22 to 30 points, with a median of 28. There were no differences in survival rate, local recurrence, or MSTS scores between patients with a tumor that did not invade the epiphysis and those in whom the tumor did invade the epiphysis. There were differences in MSTS scores among patients with epiphyseal tumor extension in which different adjuvant techniques, including microwave ablation, cryoablation, and navigation-assisted osteotomy, were used. Additional surgical procedures were recorded for 10 patients (24%). Osteonecrosis of the residual epiphysis was detected 13 patients (31%). CONCLUSIONS: Our findings suggest it is possible to salvage joints in selected patients with juxtaarticular osteosarcoma around the knee. The patients who have a favorable response to chemotherapy are the best candidates for this approach. Future studies might explore the role of adjuvant techniques of microwave ablation and cryoablation, particularly when the tumor invades the epiphysis, and whether resections can be facilitated with navigation. LEVEL OF EVIDENCE: Level IV, therapeutic study.

Individualized chemotherapy for osteosarcoma and identification of gene mutations in osteosarcoma.

The study aims to identify novel gene mutations in osteosarcoma and to guide individualized preoperative chemotherapy for osteosarcoma based on the analysis of expression and mutations of the drug-metabolism-related genes. Twenty-eight osteosarcoma patients received individualized preoperative chemotherapy regimens. Expression levels and mutations of chemotherapy-related genes in samples collected from the patients were determined using real-time PCR and DNA sequencing, respectively. Patient sensitivity to chemotherapeutic agents was evaluated by systematic analysis of the PCR and sequencing results. Novel mutations were identified via high-throughput sequencing of 339 genes in 10 osteosarcoma samples. Individualized preoperative chemotherapy outcomes were valid for nine patients (n = 9/28, 32.1%). Chemosensitivity assays showed that all 28 patients were sensitive to ifosfamide, whereas 46.4 and 39.2% were sensitive to docetaxel and platinum, respectively. More importantly, patients receiving highly chemosensitive chemotherapy agents had better prognosis and treatment outcomes than those receiving less chemosensitive agents (P < 0.05). In addition, 39 gene mutations were detected in at least five osteosarcoma tumor samples. Analysis of the expression and mutation of drug-metabolism-related genes will aid in the design of effective individualized preoperative chemotherapy regimens for osteosarcoma. Determining the chemosensitivity of individual tumors to chemotherapeutic agents will facilitate the development of better therapeutic approaches. Individualized treatment of osteosarcoma may improve chemotherapy efficacy and the survival rate of osteosarcoma patients. High-throughput genotyping allows mapping of osteosarcoma mutations, and novel gene mutations offered new candidates for diagnosis and therapeutic targeting.

Determination of cephazolin, ceftazidime, and ceftriaxone distribution in nucleus pulposus.

BACKGROUND: The intervertebral disc is the largest avascular structure in the adult body and minimal blood flow through capillary beds only supplying the outer regions of the disc, which relies on the passive diffusion as a major factor for nutrition and uptake of molecules, including antibiotics. This study is to detect the serum and nucleus pulposus (NP) levels of cephazolin, ceftazidime, and ceftriaxone and to assess this antibiotic permeability into the intervertebral disc. METHODS: Forty-five consecutive patients undergoing lumbar interbody fusion surgery were divided into three groups to participate in the study. Approximately 30 min before the procedures, a bolus dose 2 g antibiotic of cephazolin, ceftazidime, and ceftriaxone was administered intravenously. The NP tissue and serum sample levels of antibiotic were assayed by high performance liquid chromatography. RESULTS: Three cases failed in the ceftriaxone group because the NP tissue contaminates the blood. Average time between antibiotic injection and tissue/blood collection was 41 min (range 27-57 min). The antibiotic concentration level of cephazolin, ceftazidime, and ceftriaxone was 144.26 ± 29.15, 127.19 ± 30.22, and 227.81 ± 51.48 µg/ml in serum and 2.33 ± 0.45, 3.74 ± 1.91, and 2.23 ± 1.86 µg/g in NP, respectively. The antibiotic penetration in to NP of cephazolin was 1.67 ± 0.44, 2.99 ± 1.99 of ceftazidime, and 1.08 ± 1.44 of ceftriaxone. CONCLUSIONS: The antibiotics of cephazolin, ceftazidime, and ceftriaxone had concentration in the NP tissue, which was higher than the stated MIC. Ceftazidime had highest penetration in to NP tissue, and ceftriaxone had the lowest penetration in to NP tissue.

Comparative study of percutaneous vertebroplasty and kyphoplasty in the treatment of osteoporotic vertebral compression fractures.

OBJECTIVE: To compare the therapeutic effect of percutaneous vertebroplasty (PVP) and percutaneous kyphoplasty (PKP) in the treatment of osteoporotic vertebral compression fractures (VCFs). METHODS: A total of 244 patients with VCFs were treated by PVP or PKP and 192 had follow-up for at least 1 year. Clinical outcomes were determined by pain Visual Analog Scale (VAS) and Short Form 36 Health Survey (SF-36). Preoperative and postoperative radiographic assessment included measurement of posterior and anterior vertebral body height (AH and PH), as well as the kyphotic angle by the Cobb method. RESULTS: A total of 192 cases had follow-up for at least 1 year and 52 cases lost. The average amount of polymethylmethacrylate (PMMA) cement introduced per vertebra was 3.4 ± 1.5 ml in PVP and 4.5 ± 0.8 ml in PKP (P < 0.05). All patients subjectively reported immediate relief of their typical fracture pain, and the mean VAS decreased significantly from presurgery to postsurgery during the 1-year of follow-up. The RP, BP and GH dimensionality values of SF-36 in PKP were higher than PVP (P < 0.05). The improvement on AH was 11.13 ± 5.68% in PVP and 21.46 ± 9.87% in PKP (P < 0.01); on PH was 2.25 ± 1.36% in PVP and 7.57 ± 2.49% in PKP (P < 0.01). The average improvement in the kyphotic angle after the procedure was 5.21 ± 2.33° in PVP and 11.69 ± 5.18° in PKP (P < 0.01). CONCLUSIONS: PVP and PKP have the ability of reducing pain in osteoporotic VCF patients. The correction of kyphotic deformity and restoration of the anterior vertebral body heights associated with osteoporotic VCFs was better in PKP.

Percutaneous discectomy and drainage for postoperative intervertebral discitis.

BACKGROUND: Postoperative intervertebral discitis occurs following spinal surgery. This study was done to evaluate the effect of percutaneous discectomy and drainage (PDD) for postoperative intervertebral discitis. METHODS: A retrospective study of postoperative intervertebral discitis treated by PDD procedures was conducted from January 1997 to June 2006. There were 34 patients (24 males, 10 females); 10 cases of after lumbar discectomy (L3-4 in 3 patients, L4-5 in 7 patients), 21 cases of after percutaneous lumbar discectomy (L3-4 in 7 patients, L4-5 in 14 patient), 2 cases of after percutaneous cervical discectomy (C5-6 in 1 patient, C6-7 in 1 patient), and 1 case of C5-6 after percutaneous cervical nucleoplasty. RESULTS: All patients tolerated the procedure well and there were 31 cases had followed up. VAS scores demonstrated statistically significant improvement after PDD when compared with preoperational values (P < 0.01). Elevated CRP and ESR values returned to normal range within 3-8 weeks. CRP and ESR values demonstrated statistically significant improvement after PDD when compared with preoperative values (P < 0.01). Biopsies of the disc were performed in all patients and pus was seen in 17 patients at the pathology levels. Inflammatory cells were observed nine cases (4 cases showed infiltration of lymphocytes and plasmacytes, 5 cases showed infiltrate of polymorphonuclear leucocytes). Cultures of disc and bone tissue showed 17 cases of sterile and 14 had positive culture. Spine X-rays films showed narrowed disc space in 29 cases, and bridging osteophytes were noted in 19 patients. Destructive and sclerotic changes of vertebral bodies with narrowing of disc spaces were observed in 14 patients. CONCLUSIONS: The results show that PDD is a minimally invasive procedure for obtaining sufficient biopsy material for histological analysis and culture in cases of discitis, and has a good clinical outcome recommended for patients with early stage postoperative intervertebral discitis without neurologic deficit.

Percutaneous cervical nucleoplasty and percutaneous cervical discectomy treatments of the contained cervical disc herniation.

BACKGROUND: There were no studies in literature to compare the clinical outcomes of percutaneous nucleoplasty (PCN) and percutaneous cervical discectomy (PCD) in contained cervical disc herniation. METHODS: A retrospective of patients with symptomatic contained cervical disc herniated were operated on with PCN and PCD from June 2003 to July 2005. Two-hundred and four patients initially fulfilled the study criteria, and 28 patients were lost in follow-up. The patients were categorized into different groups depending on the procedure by PCN (81 cases) or PCD (95 cases). RESULTS: The clinical outcomes, pain reduction, and segment stability were recorded during this study. Puncture of the needle into the disc space was accurately performed under C-arm fluoroscopy guidance in all cases and no intraoperative deaths were reported in our study. At the end, 176 cases had follow-up and 28 cases were lost, and the follow-up rate was 88.0% (81/92) in the PCN group and 84.8% (95/112) in the PCD group. The follow-up time ranged from 16 to 48 months (average 29 months), and on an average of 28.86 ± 4.52 months on PCN and 8.42 ± 3.21 months on PCD (t = -0.24, P = 0.81, >0.05). The operation time averages of PCN and PCD are 4.67 ± 1.16 and 11.95 ± 1.80, respectively (P < 0.01). The pain index improved from 7.12 ± 1.13 to 2.74 ± 0.89 (t = 27.03, P = 0.0000, <0.001) in PCN patients and from 7.18 ± 1.09 to 2.71 ± 0.91 (t = 29.57, P = 0.0000, <0.001) in PCD patients. Clinical results of PCN were excellent in 31 cases, good 32 cases, fair 13 cases, and poor 5 cases; for PCD, the results were 33, 42, 12, and 7 cases, respectively, and 1 in discitis. Good and excellent was 78.4% (77.8% in PCN and 79.5% in PCD, P > 0.05). There was one case of PCN that had the partial Perc-D SpineWand broken in the disc space, cannot be moved by the percutaneous cervical discectomy, and remained there itself. One of the cases had discitis in this study after PCD. Patient presented with neck pain and associated radicular pain and numbness in the left upper-limb after 8 days of PCD. There were no instable cases after procedures of PCN and PCD. There were no significant difference in stability of preoperatively and postoperatively between PCN and PCD (P > 0.05). CONCLUSIONS: PCN and PCD treatments of contained cervical disc herniation show good outcomes and there was no difference in the stability of cervical spine. PCN and PCD are safe, minimally invasive, and no differences were observed between the methods in clinical outcome.