Development of betabodies: The next generation of phosphatidylserine targeting agents.

Externalized phosphatidylserine (PS) is a phospholipid and a selective marker of the tumor microenvironment (TME). It is exposed on the outer leaflet of the plasma membrane of tumor-associated endothelial cells, apoptotic tumor cells, and some viable tumor cells, where it functions in part to suppress immune responses by binding to PS receptors expressed on tumor-infiltrating myeloid cells. PS has been targeted with antibodies, such as bavituximab, that bind the phospholipid via a cofactor, ß2-glycoprotein 1 (ß2GP1); these antibodies showed excellent specificity for tumor vasculature and induce an immune stimulatory environment. We have advanced this concept by developing the next generation of PS targeting agent, a fusion protein (betabody) constructed by linking PS-binding domain V of ß2GP1 to the Fc of an IgG2a. Betabodies bind to externalized PS with high affinity (∼1 nM), without the requirement of a co-factor and localize robustly to the TME. We demonstrate that betabodies are a direct PS-targeting agent that has the potential to be used as anti-tumor therapy, drug delivery vehicles, and tools for imaging the TME.

Observation of four self-sweeping regimes in a single-mode bi-directional ytterbium-doped fiber ring laser.

In this work, for the first time, four self-sweeping regimes in a single-mode bi-directional ytterbium-doped fiber ring laser are observed by adjusting the polarization controller (PC): normal self-sweeping, reverse self-sweeping, mixed state, and wavelength stop state. In addition, regulating the PC can artificially selectively make the laser operate in normal self-sweeping or reverse self-sweeping within a certain pump power range, and their self-sweeping characteristics (e.g., sweeping rate, sweeping range, etc.) and intensity dynamics are investigated in detail, respectively. In conclusion, we can flexibly regulate the sweeping direction and sweeping characteristics of the bi-directional self-sweeping fiber ring laser in a simple approach by adjusting the PC, which is potentially valuable for its practical application.

Detection of phosphatidylserine-positive exosomes for the diagnosis of early-stage malignancies.

BACKGROUND: There has been increasing interest in the detection of tumour exosomes in blood for cancer diagnostics. Most studies have focussed on miRNA and protein signatures that are surrogate markers for specific tumour types. Because tumour cells and tumour-derived exosomes display phosphatidylserine (PS) in their outer membrane leaflet, we developed a highly sensitive ELISA-based system that detects picogram amounts of exosomal phospholipid in plasma as a cancer biomarker. METHODS: This report describes the development of a highly specific and sensitive ELISA for the capture of PS-expressing tumour exosomes in the blood of tumour-bearing mice. To monitor the relationship between tumour burden and tumour exosome plasma concentrations, plasma from one transplantable breast cancer model (MDA-MB-231) and three genetic mouse models (MMTV-PyMT; breast and KIC and KPC; pancreatic) were screened for captured exosomal phospholipid. RESULTS: We show that quantitative assessment of PS-expressing tumour exosomes detected very early-stage malignancies before clinical evidence of disease in all four model systems. Tumour exosome levels showed significant increases by day 7 after tumour implantation in the MDA-MB-231 model while palpable tumours appeared only after day 27. For the MMTV-PyMT and KIC models, tumour exosome levels increased significantly by day 49 (P⩽0.0002) and day 21 (P⩽0.001) while tumours developed only after days 60 and 40, respectively. For the KPC model, a significant increase in blood exosome levels was detected by day 70 (P=0.023) when only preinvasive lesions are microscopically detectable. CONCLUSIONS: These data indicate that blood PS exosome levels is a specific indicator of cancer and suggest that blood PS is a biomarker for early-stage malignancies.

Antibody-Mediated Blockade of Phosphatidylserine Enhances the Antitumor Effect of Sorafenib in Hepatocellular Carcinomas Xenografts.

BACKGROUND: Currently, the only FDA-approved systemic therapy for hepatocellular carcinoma (HCC) is the multi-receptor tyrosine kinase inhibitor, sorafenib, which provides only modest clinical benefit. We recently showed that treatment with a phosphatidylserine (PS) targeting agent suppresses tumor growth by targeting tumor vasculature and reactivating antitumor immunity. METHODS: We tested the hypothesis that sorafenib increases PS exposure on tumor vasculature, thereby enhancing the antitumor efficacy of PS targeting. We evaluated the efficacy of combining a PS targeting agent (2aG4) with sorafenib in murine xenograft models of human HCC. RESULTS: Our results demonstrate that combination of 2aG4 and sorafenib had a superior therapeutic effect over single agent therapy. Mechanistic studies showed that sorafenib significantly increased PS exposure on tumor vasculature; the percentage of PS-positive vessels increased from 19 to 52, 23 to 68, and 30 to 55 % in PLC/PRF/5, C3A, and Huh7 tumors, respectively. Combination therapy significantly decreased tumor microvessel density and the level of M2 macrophages, while increasing the apoptotic index of tumor endothelial cells and the frequency of M1 macrophages. Furthermore, we report the findings of a Phase I clinical study of bavituximab, a chimeric version of 2aG4, combined with sorafenib in HCC patients. The Phase I results demonstrate the appropriate dose of bavituximab to be given with sorafenib in future clinical trials. CONCLUSIONS: Overall, these results strongly support the combination of bavituximab with sorafenib as a promising systemic therapeutic strategy for the treatment for advanced HCC patients.

Molecular cloning and expression analysis of chymotrypsin-like serine protease from the redclaw crayfish (Cherax quadricarinatus): a possible role in the junior and adult innate immune systems.

A novel chymotrypsin-like serine protease (CLSP) was isolated from the hepatopancreas of the redclaw crayfish Cherax quadricarinatus (Cq-chy). The full-length cDNA of Cq-chy contains 951 nucleotides encodes a peptide of 270 amino acids. The mature peptide comprising 223 amino acids contains the conserved catalytic triad (H, D, and S). Similarity analysis showed that Cq-chy shares high identity with chymotrypsins from the fiddler crab; Uca pugilator. Cq-chy mRNA expression in C. quadricarinatus was shown to be: (a) tissue-related with the highest expression in the hepatotpancreas and widely distributed, (b) highly responsive in the hepatopancreas to White Spot Syndrome Virus (WSSV) challenge, and (c) differently regulated in immature and adult crayfish. In this study we successfully isolated Cq-chy. Our observations indicate that Cq-chy is differently involved in the immature and adult innate immune reactions, thus suggesting a role for CLSPs in the invertebrate innate immune system.

Periosteal distraction for foot preservation contributes to favorable clinical effects and prognosis in a foot ulcer patient with critical limb ischemia after multiple operations.

This paper presents the challenges faced by a patient undergoing limb-sparing treatment due to chronic limb-threatening ischemia (CLTI) complicated with long-standing nonhealing foot ulcer and intense pain. However, after multiple vascular surgeries, the foot wound continued to worsen, which could lead to transfemoral amputation and even death. We report a case of an aged male patient admitted after complaining of "pain and ulceration in his left foot for ten months". The patient was diagnosed with arteriosclerosis obliterans of the lower limbs with critical limb ischemia, which improved little after the drug therapy. This patient had undergone three endovascular procedures with a medical history of myocardial infarction and stenting. The main artery could not be directly connected to the foot by open or endovascular surgery due to severe vascular occlusion below the knee. In addition, foot ulcers made it impossible to walk, which induced angina pectoris. After coordination and discussion, we determined to perform a 2-week lateral tibial periosteum distraction (LTPD). The procedure significantly improved the foot wound and relieved the pain. After the two-week personalized wound management, the wound healed, and the pain disappeared. Consequently, the patient was able to walk independently, without recurrence during the 3-month follow-up period. Periosteal distraction has rarely been reported in previous literature and is mainly used to treat patients with diabetic foot, rather than those who have undergone repeated percutaneous transluminal angioplasty (PTA) for CLTI combined with foot ulcers. As most CLTI patients are suffering from cardiac, cerebral, and renal diseases, their blood vessels are difficult to open, with high re-occlusion and recurrence rates and low limb salvage rate. Therefore, we present our case herein so that the CLTI patients whose inferior genicular arteries cannot be opened due to severe infrapopliteal arterial occlusion complicated with nonhealing foot ulcers or intractable pain can be treated with LTPD, thus providing them with the "last kilometer" bloodstream towards the foot.

Sequential treatment for diabetic foot ulcers in dialysis patients: A case report.

BACKGROUND: Diabetic foot ulcers (DFUs) are common in patients with diabetes, especially those undergoing hemodialysis. In severe cases, these ulcers can cause damage to the lower extremities and lead to amputation. Traditional treatments such as flap transposition and transfemoral amputation are not always applicable in all cases. Therefore, there is a need for alternative treatment methods. CASE SUMMARY: This report describes a 62-year-old female patient who was admitted to the hospital with plantar and heel ulcers on her left foot. The patient had a history of renal failure and was undergoing regular hemodialysis. Digital subtraction angiography showed extensive stenosis and occlusion in the left superficial femoral artery, left peroneal artery and left posterior tibial artery. Following evaluation by a multidisciplinary team, the patient was diagnosed with type 2 DFUs (TEXAS 4D). Traditional treatments were deemed unsuitable, and the patient was treated with endovascular surgery in the affected area, in addition to supportive medical treatment, local debridement, and sequential repair using split-thickness skin and tissue-engineered skin grafts combined with negative pressure treatment. After four months, the wound had completely healed, and the patient was able to walk with a walking aid. CONCLUSION: This study demonstrates a new treatment method for DFUs was successful, using angioplasty, skin grafts, and negative pressure.

The Effect of Halo-Pelvic Traction on Bone Mineral Density of Vertebrae and Corresponding Risk Factors.

OBJECTIVES: Decreased bone mineral density (BMD) is associated with complications in implantation surgery for severe spinal deformity. In this quantitative study, we aimed to investigate the impact of halo-pelvic traction on vertebral bone mineral density (BMD) and identify the risk factors for a decrease in BMD. METHODS: Patients who underwent halo-pelvic traction at our hospital between 2019 and 2022 were included in the study. Patients' data, including height, weight, and BMD pre- and post-traction, were collected and analyzed. Quantitative computed tomography (QCT) was used to determine the BMD. The paired rank sum test was used to evaluate the changes in each measurement parameter. Linear regression was used to identify risk factors for a decrease in BMD. RESULTS: Fifteen patients were included in the study, nine women and six men, with an average age of 21.2 ± 7.3 years. Eleven patients had severe rigid scoliosis, while four had tuberculotic kyphosis. One expert measured the BMD values of 345 vertebrae using QCT. The average traction time was 143.3 ± 44.4 days. The average pre-traction BMD was 183.1 ± 73.8 mg/cm3 , and the average post-traction BMD was 140.5 ± 61.3 mg/cm3 (p < 0.01) Patients' height increased from an average of 151.3 ± 12.8 cm pre-traction to 165.5 ± 13.7 cm post-traction (p < 0.01), with traction length averaging 14.3 ± 6.2 cm (p < 0.01). The Cobb angle of the main curve declined from an average of 112.5° ± 24.4° pre-traction to 67.7° ± 19.8° post-traction (p < 0.01). Linear regression revealed a positive correlation between BMD loss and traction length and a negative correlation between BMD loss and correction rate. CONCLUSIONS: Halo-pelvic traction can lead to a decrease in the BMD of the spinal vertebrae, with traction length positively correlated with BMD loss and correction rate negatively correlated with BMD loss. To prevent osteoporosis, physicians should ensure a limited traction length while utilizing better management techniques.

The role of multi-modal intra-operative neurophysiological monitoring in corrective surgeries for thoracic tuberculosis with kyphosis.

OBJECTIVE: The aim of this study was to assess the performance and utility of motor evoked potentials (MEP) and somatosensory evoked potentials (SSEP) during corrective surgery for thoracic tuberculosis with kyphosis (TTK). METHODS: 68 patients (mean age 31.7 ± 20.3 years) who underwent corrective surgery for TTK from 2012 to 2019 were included in this retrospective study. Patients were neurologicaly evaluated before and after surgery with systematic neurologic examinations. Intraoperative neurophysiological monitoring (IONM) with SSEP and MEP was carried out. A receiver operating characteristic (ROC) curve and area under ROC curve (AUC) were used to identify the diagnostic accuracy of potential recovery. RESULTS: IONM alerting occurred in 12 surgeries (12/68, 17.6%), of which 6 were SSEP alerting, 2 MEP alerting, and 4 combinations of both SSEP and MEP. Among the 12 cases where there was IONM alerting, 3 (25%) had postoperative neurological deficits(PND), whereas one patient had PND without IONM alerting. IONM sensitivity and specificity were 0.75 (95% CI 0.22-0.99) and 0.86 (95% CI 0.74-0.93) respectively. Positive predictive value (PPV) and negative predictive value (NPV) were 0.25 and 0.98 respectively. The AUC of evoked potential recovery in diagnosing PND was 0.884. CONCLUSION: Our study showed that multi-modal IONM with SSEP and MEP can effectively indicate a potential neural injury and predict PND during TTK corrective surgery. LEVEL OF EVIDENCE: Level IV, Therapeutic Study.

Circular RNA circDNM3OS Functions as a miR-145-5p Sponge to Accelerate Cholangiocarcinoma Growth and Glutamine Metabolism by Upregulating MORC2.

BACKGROUND: Cholangiocarcinoma (CCA) is the second most common liver malignant tumor. CircRNA hsa_circ_0005230 (circDNM3OS) has been reported to exert an oncogenic role in CCA. However, the mechanisms related to circDNM3OS in CAA progression have not been fully elucidated. METHODS: The expression of circDNM3OS, microRNA (miR)-145-5p, and MORC2 (MORC Family CW-Type Zinc Finger 2) mRNA were analyzed by quantitative real-time polymerase chain reaction (qRT-PCR). Cell proliferation, colony formation, migration, invasion, and apoptosis were evaluated by Cell Counting Kit-8 (CCK-8), colony formation, transwell, wound-healing, and flow cytometry assays. The levels of glutamine, α-KG (α-ketoglutarate), and ATP (adenosine triphosphate) were detected using commercial kits. The relationship between circDNM3OS or MORC2 and miR-145-5p was verified by dual-luciferase reporter and/or RNA immunoprecipitation (RIP) assays. Protein level of MORC2 was measured by Western blotting. The role of circDNM3OS in CCA growth was verified by xenograft experiment. RESULTS: CircDNM3OS and MORC2 were upregulated while miR-145-5p was downregulated in CCA tissues and cells. Inhibition of circDNM3OS reduced xenograft tumor growth in vivo and constrained proliferation, colony formation, migration, invasion, induced apoptosis, and reduced glutamine metabolism of CCA cells in vitro. CircDNM3OS sponged miR-145-5p to elevate MORC2 expression. MiR-145-5p silencing overturned circDNM3OS knockdown-mediated influence on malignancy and glutamine metabolism of CCA cells. Also, MORC2 overexpression reversed the repressive impact of miR-145-5p mimic on malignancy and glutamine metabolism of CCA cells. CONCLUSION: CircDNM3OS facilitates CCA growth and glutamine metabolism by regulating the miR-145-5p/MORC2 pathway, offering a novel mechanism to understand the progression of CCA.