Anti-tumor effects of novel alkannin derivatives with potent selectivity on comprehensive analysis.

BACKGROUND: Therapeutic approaches based on glycolysis and energy metabolism of tumor cells are new promising strategies for the treatment of cancer. Currently, researches on the inhibition of pyruvate kinase M2, a key rate limiting enzyme in glycolysis, have been corroborated as an effective cancer therapy. Alkannin is a potent pyruvate kinase M2 inhibitor. However, its non-selective cytotoxicity has affected its subsequent clinical application. Thus, it needs to be structurally modified to develop novel derivatives with high selectivity. PURPOSE: Our study aimed to ameliorate the toxicity of alkannin through structural modification and elucidate the mechanism of the superior derivative 23 in lung cancer therapy. METHODS: On the basis of the principle of collocation, different amino acids and oxygen-containing heterocycles were introduced into the hydroxyl group of the alkannin side chain. We examined the cell viability of all derivatives on three tumor cells (HepG2, A549 and HCT116) and two normal cells (L02 and MDCK) by MTT assay. Besides, the effect of derivative 23 on the morphology of A549 cells as observed by Giemsa and DAPI staining, respectively. Flow cytometry was performed to assess the effects of derivative 23 on apoptosis and cell cycle arrest. To further assess the effect of derivative 23 on the Pyruvate kinase M2 in glycolysis, an enzyme activity assay and western blot assay were performed. Finally, in vivo the antitumor activity and safety of the derivative 23 were evaluated by using Lewis mouse lung cancer xenograft model. RESULTS: Twenty-three novel alkannin derivatives were designed and synthesized to improve the cytotoxicity selectivity. Among these derivatives, derivative 23 showed the highest cytotoxicity selectivity between cancer and normal cells. The anti-proliferative activity of derivative 23 on A549 cells (IC50 = 1.67 ± 0.34 µM) was 10-fold higher than L02 cells (IC50 = 16.77 ± 1.44 µM) and 5-fold higher than MDCK cells (IC50 = 9.23 ± 0.29 µM) respectively. Subsequently, fluorescent staining and flow cytometric analysis showed that derivative 23 was able to induce apoptosis of A549 cells and arrest the cell cycle in the G0/G1 phase. In addition, the mechanistic studies suggested derivative 23 was an inhibitor of pyruvate kinase; it could regulate glycolysis by inhibiting the activation of the phosphorylation of PKM2/STAT3 signaling pathway. Furthermore, studies in vivo demonstrated derivative 23 significantly inhibited the growth of xenograft tumor. CONCLUSION: In this study, alkannin selectivity is reported to be significantly improved following structural modification, and derivative 23 is first shown to be able to inhibit lung cancer growth via the PKM2/STAT3 phosphorylation signaling pathway in vitro, indicating the potential value of derivative 23 in treating lung cancer.

The Dual Benefit of Sulfasalazine on Pneumocystis Pneumonia-Related Immunopathogenesis and Antifungal Host Defense Does Not Require IL-4Rα-Dependent Macrophage Polarization.

Pneumocystis is a respiratory fungal pathogen that is among the most frequent causes of life-threatening pneumonia (PcP) in immunocompromised hosts. Alveolar macrophages play an important role in host defense against Pneumocystis, and several studies have suggested that M2 polarized macrophages have anti-Pneumocystis effector activity. Our prior work found that the immunomodulatory drug sulfasalazine (SSZ) provides a dual benefit during PcP-related immune reconstitution inflammatory syndrome (IRIS) by concurrently suppressing immunopathogenesis while also accelerating macrophage-mediated fungal clearance. The benefits of SSZ were associated with heightened Th2 cytokine production and M2 macrophage polarization. Therefore, to determine whether SSZ improves the outcome of PcP through a mechanism that requires Th2-dependent M2 polarization, RAG2-/- mice lacking interleukin 4 receptor alpha chain (IL-4Rα) on macrophage lineage cells were generated. As expected, SSZ treatment dramatically reduced the severity of PcP-related immunopathogenesis and accelerated fungal clearance in immune-reconstituted RAG2-/- mice. Similarly, SSZ treatment was also highly effective in immune-reconstituted RAG2/IL-4Rα-/- and RAG2/gamma interferon receptor (IFN-γR)-/- mice, demonstrating that neither IL-4Rα-dependent M2 nor IFN-γR-dependent M1 macrophage polarization programs were required for the beneficial effects of SSZ. Despite the fact that macrophages from RAG2/IL-4Rα-/- mice could not respond to the Th2 cytokines IL-4 and IL-13, M2-biased alveolar macrophages were identified in the lungs following SSZ treatment. These data demonstrate that not only does SSZ enhance phagocytosis and fungal clearance in the absence of macrophage IL-4Rα signaling, but also that SSZ promotes M2 macrophage polarization in an IL-4Rα-independent manner. These findings could have implications for the treatment of PcP and other diseases in which M2 polarization is beneficial.

Ultrasound-Guided Clavipectoral Fascial Plane Block With Intermediate Cervical Plexus Block for Midshaft Clavicular Surgery: A Prospective Randomized Controlled Trial.

BACKGROUND: Regional anesthesia such as interscalene brachial plexus block (ISBPB) with intermediate cervical plexus block (ICPB) is generally a preferred choice for clavicular surgery. However, various studies have shown that these blocks, especially ISBPB, could cause phrenic nerve paralysis and decrease diaphragmatic motion. The study aimed to evaluate the efficacy of clavipectoral fascial plane block (CPB), an alternative technique to ISBPB, with ICPB, in reducing hemidiaphragmatic paralysis during midshaft clavicular surgery. METHODS: Forty patients scheduled for right midshaft clavicular surgery were randomized (1:1) into an ultrasound-guided ISBPB with ICPB (BC) group or ultrasound-guided CPB with ICPB (CC) group. Five milliliter of 0.375% ropivacaine was used for ICPB, another 20 mL for ISBPB or CPB, and no administration of additional sedative or general anesthetic was planned. Primary outcome was measured by the incidence of hemidiaphragmatic paralysis using M-mode ultrasonography, while secondary outcomes were measured by bedside pulmonary function test, the success rate of block, the time required for the block procedure and onset of block, and motor block score in right upper extremity. RESULTS: In comparison with BC group, the incidence of hemidiaphragmatic paralysis postblock was decreased in CC group (50% vs 0%; P < .001), and measurement of bedside pulmonary function was significantly improved. There was a 100% success rate for anesthetic block in both BC and CC groups, and CC group showed lower motor block score in upper extremity and less block procedure time than BC group (7.1 ± 1.2 vs 3.2 ± 0.6 minutes; P < .001). Moreover, no significant differences were found between time of onset of block and other anesthetic complications in the 2 groups. CONCLUSIONS: Ultrasound-guided CPB with ICPB could significantly reduce hemidiaphragmatic paralysis and provide an adequate surgical anesthesia with fewer complications such as motor block in upper extremity during right midshaft clavicular surgery.

Complex renal calculi treated with traditional Chinese medicine Paishi decoction combined with laparoscopic ureterectomy.

To analyze the effect of traditional Chinese medicine Paishi decoction combined with laparoscopic ureterectomy and lithotripsy in the treatment of complex kidney stones. Totally 100 patients with complicated kidney stones admitted to our hospital from January 2019 to January 2021 were selected and randomly divided into a control group and an experimental group, with 50 cases in each group. The control group was treated with laparoscopic ureterectomy for stone removal, the experimental group was treated with traditional Chinese medicine Paishi decoction combined with laparoscopic ureterectomy for stone removal. The therapeutic effects of the two groups were compared. The total effective rate of treatment in the control group was 76% and that of the experimental group was 96%. The stone clearing time, time to pain resolution and time to hematuria disappearance time in the experimental group were significantly shorter as compared with the control group. After treatment, the levels of serum creatinine and blood urea nitrogen in the experimental group were significantly lower than those in the control group. Traditional Chinese medicine Paishi decoction combined with laparoscopic ureterectomy and lithotripsy for treatment of complex kidney stones ameliorates the treatment efficacy, shortens the time of stone removal, mitigates the clinical symptoms of patients, and helps restore renal function, which is worthy of clinical promotion and application.

Lysates of Lactobacillus acidophilus combined with CTLA-4-blocking antibodies enhance antitumor immunity in a mouse colon cancer model.

Previous reports have suggested that many gut microbiomes were associated with the development of colorectal cancer (CRC), and could modulate response to numerous forms of cancer therapy, including checkpoint blockade immunotherapy. Here we evaluated the protective efficacy of Lactobacillus acidophilus (L. acidophilus) cell lysates combined with an anti-CTL antigen-4 blocking antibody (CTLA-4 mAb) in syngeneic BALB/c mice CRC models induce by a single intraperitoneal injection of 10 mg/kg azoxymethane (AOM), followed by three cycles of 2% dextran sulfate sodium (DSS) in drinking water. In contrast to CTLA-4 mAb monotherapy, L. acidophilus lysates could attenuate the loss of body weight and the combined administration significantly protected mice against CRC development, which suggested that the lysates enhanced antitumor activity of CTLA-4 mAb in model mice. The enhanced efficacy was associated with the increased CD8 + T cell, increased effector memory T cells (CD44 + CD8 + CD62L+), decreased Treg (CD4 + CD25 + Foxp3+) and M2 macrophages (F4/80 + CD206+) in the tumor microenvironment. In addition, our results revealed that L. acidophilus lysates had an immunomodulatory effect through inhibition the M2 polarization and the IL-10 expressed levels of LPS-activated Raw264.7 macrophages. Finally, the 16S rRNA gene sequencing of fecal microbiota demonstrated that the combined administration significantly inhibited the abnormal increase in the relative abundance of proteobacteria and partly counterbalance CRC-induced dysbiosis in model mice. Overall, these data support promising clinical possibilities of L. acidophilus lysates with CTLA-4 mAb in cancer patients and the hypothesis that probiotics help shape the anticancer immune response.

Malignant ascite-derived extracellular vesicles inhibit T cell activity by upregulating Siglec-10 expression.

Background and purpose: To evade immune defense, cancer cells can employ extracellular vesicles (EVs) to inhibit the anti-tumor activity of lymphocytes in the tumor microenvironment. However, the mechanisms and key molecules that mediate the effects of EVs on lymphocytes are unclear. Patients and methods: We used Quantibody® Human Cytokine Antibody Array 440 to determine the tumor immunity-related cytokine profile of peripheral blood lymphocytes (PBLs) stimulated with EVs derived from peritoneal washes or malignant ascites. We detected 21 upregulated and 27 downregulated proteins, including the immunosuppressive receptors Siglec-10, SLAM, PD-1, and TIM-3. Results: Flow cytometry analysis of PBLs or ovarian cancer ascites suggested that Siglec-10 expression on CD3+ T cells was higher in ovarian cancer patients than in healthy controls and in the malignant ascites of ovarian cancer patients than in their blood. Moreover, the expression of CD24, the Siglec-10 ligand, was associated with tumor stage and cancer cell metastasis. Finally, compared to the benign peritoneal wash-derived EVs, the malignant EVs significantly upregulated Siglec-10 expression on Jurkat T cells, inhibited the protein kinase C activity induced by phorbol 12-myristate 13-acetate and ionomycin, and impaired the phosphorylation of the tyrosine kinase ZAP-70 activated by crosslinking with an anti-CD3 antibody. Conclusion: The EVs secreted by malignant ovarian cells upregulated Siglec-10 expression on T cells and impaired T cell activation in the tumor microenvironment. We believe that a comprehensive understanding of the regulation of Siglec-10 and CD24 by malignant EVs has clinical importance, as it will aid in the development of better immunotherapeutic strategies for ovarian cancer.

Effect of iron nanoparticles on passivation of cadmium in the pig manure aerobic composting process.

Cadmium (Cd) is a toxic metal ion in pig manure impacting on the ecosystem, and hence the immobilization of Cd by green synthesis of iron nanoparticles (G-nFe) is a potential approach. In this study, transformation of Cd (II) during the pig manure thermophilic aerobic composting process in the presence of G-nFe was investigated. The results show that the addition of G-nFe promoted the composting process and release of available phosphorus (AP). In all six experiments, obvious passivation of Cd occurred during 15 days' composting. Particularly when 500 mL kg-1 of G-nFe was added and Cd (II) was added at 0.6%(w/w%), residual Cd increased from 0.0016% to 55.70% and exchangeable Cd decreased from 98.54% to 7.21%. Batch experiments revealed that the G-nFe promoted the transformation of Cd into a larger passivation fraction. X-ray photoelectron spectroscopy (XPS), X-ray diffraction (XRD), SEM-Mapping and Fourier transform infrared (FTIR) analysis was used to characterize residual samples, where indicated that the passivation of Cd in compost was highly correlated with the increase of P, it can be concluded that fixing with compost resulted in the formation of Cd phosphate precipitation or co-precipitation with other phosphates.

Effect of Shenmai injection on cognitive function after cardiopulmonary bypass in cardiac surgical patients: a randomized controlled trial.

BACKGROUND: Postoperative cognitive dysfunction (POCD) is a common complication after cardiac surgery that influences the clinical outcomes and quality of life of patients. This study aimed to evaluate the effects of Shenmai injection (SMI) on POCD of patients who underwent cardiac valve replacement under cardiopulmonary bypass (CPB). METHODS: This prospective, randomized, controlled trial was conducted from September 2014 to January 2017. Eighty-eight patients receiving cardiac valve replacement under CPB were randomized into the control (C) or the SMI (S) group. SMI (0.6 mL/kg) was administered intravenously from the time of anesthesia induction to the beginning of CPB. Cognitive function was assessed at 3 days before surgery and 3 days, 7 days, and 1 month after surgery using the Beijing version of the Montreal Cognitive Assessment (MoCA-BJ) score. The serum levels of neuroglobin (Ngb), hypoxia-inducible factor-1α (HIF-1α), and neuron-specific enolase (NSE) were measured at 30 min after induction (T0), immediately after the endonasal temperature rewarmed to 36 °C (T1), and 1 h (T2), 6 h (T3), 24 h (T4), 48 h (T5), and 72 h (T6) after CPB. RESULTS: Compared with the baseline values at T0, the serum Ngb levels in group C were significantly decreased at T1-2 and then increased at T3-6, while the levels in group S were decreased at T1-2 and increased at T4-6, compared to group C (p < 0.05). The serum HIF-1α levels at T1-4 and the serum NSE levels at T1-6 were significantly increased in both groups (p < 0.05). The serum levels of Ngb at T3, HIF-1α at T1-3, and NSE at T3-4,6 were lower in group S, compared to group C (p < 0.01). The MoCA-BJ scores were decreased at 3 and 7 days after surgery in both groups, and the MoCA-BJ scores in group S were higher than those in group C at 3 and 7 days after surgery (p < 0.01). CONCLUSION: Cognitive function is impaired postoperatively in patients who have undergone cardiac valve replacement under CPB. In addition, treatment with the traditional Chinese medicine SMI decreases the serum levels of Ngb, HIF-1α, and NSE as well as attenuates cognitive dysfunction. TRIAL REGISTRATION: This trial was registered with Clinicaltrials.gov as ChiCTR-TRC-14004373 on March 11, 2014.

Knockdown of EPCR inhibits the proliferation and migration of human gastric cancer cells via the ERK1/2 pathway in a PAR-1-dependent manner.

Endothelial protein C receptor (EPCR) has been implicated in the carcinogenesis of diverse tumor types. This tumor-promoting effect of EPCR is associated with the upregulation of activated protein C and the activation of protease-activated receptor 1 (PAR-1). However, the exact role of EPCR in gastric cancer (GC) and the mechanisms underlying the regulation of EPCR remain elusive. In the present study, we investigated the effects of EPCR on human GC cells, as well as the underlying mechanisms. An siRNA inference system was used to knock down the expression of EPCR in GC cells, and CCK-8, colony formation and Transwell assays were performed to determine the effects of EPCR knockdown on the proliferation and migration of the tumor cells. Additionally, cell cycle distribution and apoptosis were assessed by flow cytometry, and activated PAR-1 levels were determined by cell ELISA. The results indicated that the proliferation, clonogenicity and migration were significantly reduced and that the cell cycle was arrested in the Gap 1 phase by EPCR knockdown in SGC7901 and AGS cells. Meanwhile, apoptosis was promoted by EPCR knockdown in the two cell lines. The activation of PAR-1 on the cell surface of SGC7901 and AGS cells was significantly reduced after the knockdown of EPCR. By contrast, blockade of PAR-1 reduced the proliferation and migration of gastric cells in vitro. Additionally, after the knockdown of EPCR or treatment with PAR-1 antibody, the expression of pERK1/2 was significantly downregulated in the SGC7901 and AGS cells, while the expression levels of p-AKT (S473) and p-AKT (T308) were unchanged. The findings of the present study demonstrated that EPCR exerts pro-carcinogenic effects in GC cells in a PAR-1-dependent manner via the ERK1/2-MAPK pathway. Thus, EPCR may be a potential molecular diagnostic or therapeutic target for GC.

COX-2 inhibition attenuates lung injury induced by skeletal muscle ischemia reperfusion in rats.

BACKGROUND: Skeletal muscle ischemia reperfusion accounts for high morbidity and mortality, and cyclooxygenase (COX)-2 is implicated in causing muscle damage. Downregulation of aquaporin-1 (AQP-1) transmembrane protein is implicated in skeletal muscle ischemia reperfusion induced remote lung injury. The expression of COX-2 in lung tissue and the effect of COX-2 inhibition on AQP-1 expression and lung injury during skeletal muscle ischemia reperfusion are not known. We investigated the role of COX-2 in lung injury induced by skeletal muscle ischemia reperfusion in rats and evaluated the effects of NS-398, a specific COX-2 inhibitor. METHODS: Twenty-four Sprague Dawley rats were randomized into 4 groups: sham group (SM group), sham+NS-398 group (SN group), ischemia reperfusion group (IR group) and ischemia reperfusion+NS-398 group (IN group). Rats in the IR and IN groups were subjected to 3h of bilateral ischemia followed by 6h of reperfusion in hindlimbs, and intravenous NS-398 8 mg/kg was administered in the IN group. In the SM and SN groups, rubber bands were in place without inflation. At the end of reperfusion, myeloperoxidase (MPO) activity, COX-2 and AQP-1 protein expression in lung tissue, PGE2 metabolite (PGEM), tumor necrosis factor (TNF)-α and interleukin (IL)-1ß levels in bronchoalveolar lavage (BAL) fluid were assessed. Histological changes in lung and muscle tissues and wet/dry (W/D) ratio were also evaluated. RESULTS: MPO activity, COX-2 expression, W/D ratio in lung tissue, and PGEM, TNF-α and IL-1ß levels in BAL fluid were significantly increased, while AQP-1 protein expression downregulated in the IR group as compared to that in the SM group (P<0.05). These changes were remarkably mitigated in the IN group (P<0.05). NS-398 treatment also alleviated histological signs of lung and skeletal muscle injury. CONCLUSION: COX-2 protein expression was upregulated in lung tissue in response to skeletal muscle ischemia reperfusion. COX-2 inhibition may modulate pulmonary AQP-1 expression and attenuate lung injury.

Effect of Shenmai injection, a traditional Chinese medicine, on pulmonary dysfunction after tourniquet-induced limb ischemia-reperfusion.

BACKGROUND: Tourniquet has been considered as a recognized cause of lower limb ischemia-reperfusion injury in the orthopedic field. This study investigates pulmonary function after tourniquet deflation and the protective effect of Shenmai injection (SMI), a traditional Chinese medicine. METHODS: Twenty-eight patients undergoing lower extremity surgery were randomized into a control group (group C) and a SMI group (group S), 14 patients in each group. Blood gas and circulating indicators (malondialdehyde, interleukin [IL]-6, and IL-8) were measured immediately before tourniquet inflation and at 0.5 hour, 2 hours, 6 hours, and 24 hours after tourniquet deflation. RESULTS: Plasma levels of malondialdehyde, IL-6, and IL-8 in group C were significantly increased over baselines from 2 hours to 24 hours after tourniquet deflation and the levels reached their peaks at 6 hours after tourniquet deflation, when arterial partial pressures of oxygen and arterial-alveolar oxygen tension ratio were decreased, whereas alveolar-arterial oxygen difference was increased significantly. Both the changes in blood gas variables and plasma mediators were attenuated in group S. CONCLUSION: Pulmonary gas exchange is impaired after lower limb ischemia-reperfusion induced by clinical tourniquet application. Pretreatment with SMI, a traditional Chinese medicine, attenuates lipid peroxidation and systemic inflammatory response and mitigates pulmonary dysfunction.