A Lipid Emulsion Reverses Toxic-Dose Bupivacaine-Induced Vasodilation during Tyrosine Phosphorylation-Evoked Contraction in Isolated Rat Aortae.

The goal of this in vitro study was to examine the effect of a lipid emulsion on toxic-dose bupivacaine-induced vasodilation in a model of tyrosine phosphatase inhibitor sodium orthovanadate-induced contraction in endothelium-denuded rat aortae and to elucidate the associated cellular mechanism. The effect of a lipid emulsion on vasodilation induced by a toxic dose of a local anesthetic during sodium orthovanadate-induced contraction was examined. In addition, the effects of various inhibitors, either bupivacaine alone or a lipid emulsion plus bupivacaine, on protein kinase phosphorylation induced by sodium orthovanadate in rat aortic vascular smooth muscle cells was examined. A lipid emulsion reversed the vasodilation induced by bupivacaine during sodium orthovanadate-induced contraction. The lipid emulsion attenuated the bupivacaine-mediated inhibition of the sodium orthovanadate-induced phosphorylation of protein tyrosine, c-Jun NH2-terminal kinase (JNK), myosin phosphatase target subunit 1 (MYPT1), phospholipase C (PLC) γ-1 and extracellular signal-regulated kinase (ERK). These results suggest that a lipid emulsion reverses toxic-dose bupivacaine-induced vasodilation during sodium orthovanadate-induced contraction via the activation of a pathway involving either tyrosine kinase, JNK, Rho-kinase and MYPT1 or tyrosine kinase, PLC γ-1 and ERK, and this reversal is associated with the lipid solubility of the local anesthetic and the induction of calcium sensitization.

Mepivacaine attenuates vasodilation induced by ATP-sensitive potassium channels in rat aorta.

The goal of this in vitro study was to investigate the effect of mepivacaine on vasodilation induced by the ATP-sensitive potassium (KATP) channel opener levcromakalim in isolated endothelium-denuded rat aortas. The effects of mepivacaine and the KATP channel inhibitor glibenclamide, alone or in combination, on levcromakalim-induced vasodilation were assessed in the isolated aortas. The effects of mepivacaine or combined treatment with a protein kinase C (PKC) inhibitor, GF109203X, and mepivacaine on this vasodilation were also investigated. Levcromakalim concentration-response curves were generated for isolated aortas precontracted with phenylephrine or a PKC activator, phorbol 12,13-dibutyrate (PDBu). Further, the effects of mepivacaine and glibenclamide on levcromakalim-induced hyperpolarization were assessed in rat aortic vascular smooth muscle cells. Mepivacaine attenuated levcromakalim-induced vasodilation, whereas it had no effect on this vasodilation in isolated aortas pretreated with glibenclamide. Combined treatment with GF109203X and mepivacaine enhanced levcromakalim-induced vasodilation compared with pretreatment with mepivacaine alone. This vasodilation was attenuated in aortas precontracted with PDBu compared with those precontracted with phenylephrine. Mepivacaine and glibenclamide, alone or in combination, attenuated levcromakalim-induced membrane hyperpolarization. Taken together, these results suggest that mepivacaine attenuates vasodilation induced by KATP channels, which appears to be partly mediated by PKC.

Dexmedetomidine-Induced Contraction in the Isolated Endothelium-Denuded Rat Aorta Involves PKC-δ-mediated JNK Phosphorylation.

Vasoconstriction mediated by the highly selective alpha-2 adrenoceptor agonist dexmedetomidine leads to transiently increased blood pressure and severe hypertension. The dexmedetomidine-induced contraction involves the protein kinase C (PKC)-mediated pathway. However, the main PKC isoform involved in the dexmedetomidine-induced contraction remains unknown. The goal of this in vitro study was to examine the specific PKC isoform that contributes to the dexmedetomidine-induced contraction in the isolated rat aorta. The endothelium-denuded rat aorta was suspended for isometric tension recording. Dexmedetomidine dose-response curves were generated in the presence or absence of the following inhibitors: the pan-PKC inhibitor, chelerythrine; the PKC-α and -ß inhibitor, Go6976; the PKC-α inhibitor, safingol; the PKC-ß inhibitor, ruboxistaurin; the PKC-δ inhibitor, rottlerin; the c-Jun NH2-terminal kinase (JNK) inhibitor, SP600125; and the myosin light chain kinase inhibitor, ML-7 hydrochloride. Western blot analysis was used to examine the effect of rottlerin on dexmedetomidine-induced PKC-δ expression and JNK phosphorylation in rat aortic vascular smooth muscle cells (VSMCs) and to investigate the effect of dexmedetomidine on PKC-δ expression in VSMCs transfected with PKC-δ small interfering RNA (siRNA) or control siRNA. Chelerythrine as well as SP600125 and ML-7 hydrochloride attenuated the dexmedetomidine-induced contraction. Go6976, safingol, and ruboxistaurin had no effect on the dexmedetomidine-induced contraction, whereas rottlerin inhibited the dexmedetomidine-induced contraction. Dexmedetomidine induced PKC-δ expression, whereas rottlerin and PKC-δ siRNA transfection inhibited dexmedetomidine-induced PKC-δ expression. Dexmedetomidine also induced JNK phosphorylation, which was inhibited by rottlerin. Taken together, these results suggest that the dexmedetomidine-induced contraction involves PKC-δ-dependent JNK phosphorylation in the isolated rat aorta.

Mepivacaine-induced contraction involves phosphorylation of extracellular signal-regulated kinase through activation of the lipoxygenase pathway in isolated rat aortic smooth muscle.

Mepivacaine is an aminoamide local anesthetic with an intermediate duration that intrinsically produces vasoconstriction both in vivo and in vitro. This study investigated the arachidonic acid metabolic pathways involved in mepivacaine-induced contraction, and elucidated the associated cellular mechanism with a particular focus on extracellular signal-regulated kinase (ERK) in endothelium-denuded rat aorta. Isolated rat thoracic aortic rings were suspended for isometric tension recording. Cumulative mepivacaine concentration-response curves were generated in the presence or absence of the following inhibitors: quinacrine dihydrochloride, nordihydroguaiaretic acid, phenidone, AA-861, indomethacin, NS-398, SC-560, fluconazole, PD 98059, and verapamil. Mepivacaine-induced ERK phosphorylation, 5-lipoxygenase (5-LOX) expression, and cyclooxygenase (COX)-2 expression in rat aortic smooth muscle cells were detected by Western blot analysis in the presence or absence of inhibitors. Mepivacaine produced tonic contraction in isolated endothelium-denuded rat aorta. Quinacrine dihydrochloride, nordihydroguaiaretic acid, phenidone, AA-861, NS-398, PD 98059, and verapamil attenuated mepivacaine-induced contraction in a concentration-dependent manner. However, fluconazole had no effect on mepivacaine-induced contraction. PD 98059, quinacrine dihydrochloride, nordihydroguaiaretic acid, AA-861, phenidone, and indomethacin attenuated mepivacaine-induced ERK phosphorylation. Mepivacaine upregulated 5-LOX and COX-2 expression. These results suggest that mepivacaine-induced contraction involves ERK activation, which is primarily mediated by the 5-LOX pathway and in part by the COX-2 pathway.

Vasoconstriction potency induced by aminoamide local anesthetics correlates with lipid solubility.

Aminoamide local anesthetics induce vasoconstriction in vivo and in vitro. The goals of this in vitro study were to investigate the potency of local anesthetic-induced vasoconstriction and to identify the physicochemical property (octanol/buffer partition coefficient, pKa, molecular weight, or potency) of local anesthetics that determines their potency in inducing isolated rat aortic ring contraction. Cumulative concentration-response curves to local anesthetics (levobupivacaine, ropivacaine, lidocaine, and mepivacaine) were obtained from isolated rat aorta. Regression analyses were performed to determine the relationship between the reported physicochemical properties of local anesthetics and the local anesthetic concentration that produced 50% (ED(50)) of the local anesthetic-induced maximum vasoconstriction. We determined the order of potency (ED(50)) of vasoconstriction among local anesthetics to be levobupivacaine > ropivacaine > lidocaine > mepivacaine. The relative importance of the independent variables that affect the vasoconstriction potency is octanol/buffer partition coefficient > potency > pKa > molecular weight. The ED(50) in endothelium-denuded aorta negatively correlated with the octanol/buffer partition coefficient of local anesthetics (r(2) = 0.9563; P < 0.001). The potency of the vasoconstriction in the endothelium-denuded aorta induced by local anesthetics is determined primarily by lipid solubility and, in part, by other physicochemical properties including potency and pKa.

Mitigation of H2O2-induced autophagic cell death by propofol in H9c2 cardiomyocytes.

Autophagy, a self-eating process, is responsible for degradation of long-lived proteins and damaged cellular proteins/organelles. Double-membrane autophagosomes, formed during the process, engulf proteins/organelles and fuse with lysosomes to degrade the contents. It is important to maintain cell homeostasis and many physiological processes including cellular responses to oxidative stress. Oxidative stress induced by myocardial infarction is a major factor of heart failures. In this study, we examined how propofol modulates hydrogen peroxide (H(2)O(2))-induced autophagic cell death in H9c2 cardiomyocytes. H(2)O(2) dramatically induced cell death, which was similarly reduced in the presence of either propofol or autophagy inhibitors (e.g., wortmannin), suggesting that propofol has a protective effect in H(2)O(2)-induced autophagic cell death. Acidic autophagic vacuoles were elevated in H(2)O(2)-treated H9c2 cells, but they were largely decreased in the presence of propofol. Furthermore, many autophagy-related proteins such as LC3-II, ATG proteins, p62, AMPK, and JNK were activated in H(2)O(2)-treated H9c2 cells and were significantly deactivated in the presence of propofol. These results show that propofol regulates oxidative stress-induced autophagic cell death in cardiomyocytes. We further suggest that propofol can act as a cardioprotectant in heart diseases.

Mepivacaine-induced contraction is attenuated by endothelial nitric oxide release in isolated rat aorta.

Mepivacaine is an aminoamide-linked local anesthetic with an intermediate duration that intrinsically produces vasoconstriction both in vivo and in vitro. The aims of this in-vitro study were to examine the direct effect of mepivacaine in isolated rat aortic rings and to determine the associated cellular mechanism with a particular focus on endothelium-derived vasodilators, which modulate vascular tone. In the aortic rings with or without endothelium, cumulative mepivacaine concentration-response curves were generated in the presence or absence of the following antagonists: N(ω)-nitro-L-arginine methyl ester [L-NAME], indomethacin, fluconazole, methylene blue, 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one [ODQ], verapamil, and calcium-free Krebs solution. Mepivacaine produced vasoconstriction at low concentrations (1 × 10(-3) and 3 × 10(-3) mol/L) followed by vasodilation at a high concentration (1 × 10(-2) mol/L). The mepivacaine-induced contraction was higher in endothelium-denuded aortae than in endothelium-intact aortae. Pretreatment with L-NAME, ODQ, and methylene blue enhanced mepivacaine-induced contraction in the endothelium-intact rings, whereas fluconazole had no effect. Indomethacin slightly attenuated mepivacaine-induced contraction, whereas verapamil and calcium-free Krebs solution more strongly attenuated this contraction. The vasoconstriction induced by mepivacaine is attenuated mainly by the endothelial nitric oxide - cyclic guanosine monophosphate pathway. In addition, mepivacaine-induced contraction involves cyclooxygenase pathway activation and extracellular calcium influx via voltage-operated calcium channels.

Lipid emulsion reverses Levobupivacaine-induced responses in isolated rat aortic vessels.

BACKGROUND: The goal of this in vitro study was to investigate the effects of lipid emulsion (LE) on local anesthetic levobupivacaine-induced responses in isolated rat aorta and to determine whether the effect of LE is related to the lipid solubility of local anesthetics. METHODS: Isolated rat aortic rings were suspended for isometric tension recording. The effects of LE were determined during levobupivacaine-, ropivacaine-, and mepivacaine-induced responses. Endothelial nitric oxide synthase and caveolin-1 phosphorylation was measured in human umbilical vein endothelial cells treated with levobupivacaine alone and with the addition of LE. RESULTS: Levobupivacaine produced vasoconstriction at lower, and vasodilation at higher, concentrations, and both were significantly reversed by treatment with LE. Levobupivacaine and ropivacaine inhibited the high potassium chloride-mediated contraction, which was restored by LE. The magnitude of LE-mediated reversal was greater with levobupivacaine treatment than with ropivacaine, whereas this reversal was not observed in mepivacaine-induced responses. In LE-pretreated rings, low-dose levobupivacaine- and ropivacaine-induced contraction was attenuated, whereas low-dose mepivacaine-induced contraction was not significantly altered. Treatment with LE also inhibited the phosphorylation of endothelial nitric oxide synthase induced by levobupivacaine in human umbilical vein endothelial cells. CONCLUSIONS: These results indicate that reversal of levobupivacaine-induced vasodilation by LE is mediated mainly through the attenuation of levobupivacaine-mediated inhibition of L-type calcium channel-dependent contraction and, in part, by inhibition of levobupivacaine-induced nitric oxide release. LE-mediated reversal of responses induced by local anesthetics may be related to their lipid solubility.

Calcium sensitization involved in dexmedetomidine-induced contraction of isolated rat aorta.

Dexmedetomidine, a full agonist of the α2B-adrenoceptor that is mainly involved in vascular smooth muscle contraction, is primarily used for analgesia and sedation in intensive care units. High-dose dexmedetomidine produces hypertension in children and adults. The goal of this in vitro study was to investigate the role of the calcium (Ca(2+)) sensitization mechanism involving Rho-kinase, protein kinase C (PKC), and phosphoinositide 3-kinase (PI3-K) in mediating contraction of isolated rat aortic smooth muscle in response to dexmedetomidine. The effect of dexmedetomidine on the intracellular Ca(2+) level ([Ca(2+)]i) and tension was measured simultaneously. Dexmedetomidine concentration-response curves were generated in the presence or absence of the following antagonists: rauwolscine, Y 27632, LY 294002, GF 109203X, and verapamil. Dexmedetomidine-induced phosphorylation of PKC and membrane translocation of Rho-kinase were detected with Western blotting. Rauwolscine, Y 27632, GF 109203X, LY 294002, and verapamil attenuated dexmedetomidine-induced contraction. The slope of the [Ca(2+)]i-tension curve for dexmedetomidine was higher than that for KCl. Dexmedetomidine induced phosphorylation of PKC and membrane translocation of Rho-kinase. These results suggest that dexmedetomidine-induced contraction involves a Ca(2+) sensitization mechanism mediated by Rho-kinase, PKC, and PI3-K that is secondary to α2-adrenoceptor stimulation in rat aortic smooth muscle.

Management of perioperative acute massive pulmonary embolism: A case series.

The management of acute massive pulmonary embolism during the perioperative period is challenging. Accurate diagnosis using echocardiography and application of rapid extracorporeal membrane oxygenation can improve patients' outcomes.

The proper concentrations of dextrose and lidocaine in regenerative injection therapy: in vitro study.

BACKGROUND: Prolotherapy is a proliferation therapy as an alternative medicine. A combination of dextrose solution and lidocaine is usually used in prolotherapy. The concentrations of dextrose and lidocaine used in the clinical field are very high (dextrose 10%-25%, lidocaine 0.075%-1%). Several studies show about 1% dextrose and more than 0.2% lidocaine induced cell death in various cell types. We investigated the effects of low concentrations of dextrose and lidocaine in fibroblasts and suggest the optimal range of concentrations of dextrose and lidocaine in prolotherapy. METHODS: Various concentrations of dextrose and lidocaine were treated in NIH-3T3. Viability was examined with trypan blue exclusion assay and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. Migration assay was performed for measuring the motile activity. Extracellular signal-regulated kinase (Erk) activation and protein expression of collagen I and α-smooth muscle actin (α-SMA) were determined with western blot analysis. RESULTS: The cell viability was decreased in concentrations of more than 5% dextrose and 0.1% lidocaine. However, in the concentrations 1% dextrose (D1) and 0.01% lidocaine (L0.01), fibroblasts proliferated mildly. The ability of migration in fibroblast was increased in the D1, L0.01, and D1 + L0.01 groups sequentially. D1 and L0.01 increased Erk activation and the expression of collagen I and α-SMA and D1 + L0.01 further increased. The inhibition of Erk activation suppressed fibroblast proliferation and the synthesis of collagen I. CONCLUSIONS: D1, L0.01, and the combination of D1 and L0.01 induced fibroblast proliferation and increased collagen I synthesis via Erk activation.

Lipid emulsion attenuates extrinsic apoptosis induced by amlodipine toxicity in rat cardiomyoblasts.

Amlodipine-induced toxicity has detrimental effects on cardiac cells. The aim of this study was to examine the effect of lipid emulsion on decreased H9c2 rat cardiomyoblast viability induced by amlodipine toxicity. The effects of amlodipine, lipid emulsion, LY 294002, and glibenclamide, either alone or in combination, on cell viability and count, apoptosis, and expression of cleaved caspase-3 and -8, and Bax were examined. LY 294002 and glibenclamide partially reversed lipid emulsion-mediated attenuation of decreased cell viability and count induced by amlodipine. Amlodipine increased caspase-3 and -8 expression, but it did not alter Bax expression. LY 294002 and glibenclamide reversed lipid emulsion-mediated inhibition of cleaved caspase-3 and -8 expression induced by amlodipine. Lipid emulsion inhibited early and late apoptosis induced by amlodipine. LY 294002 and glibenclamide inhibited lipid emulsion-mediated inhibition of late apoptosis induced by amlodipine, but they did not significantly alter lipid emulsion-mediated inhibition of early apoptosis induced by amlodipine. Lipid emulsion decreased amlodipine-induced TUNEL-positive cells. These results suggest that lipid emulsion inhibits late apoptosis induced by amlodipine at toxic dose via the activation of phosphoinositide-3 kinase and ATP-sensitive potassium channels in the extrinsic apoptotic pathway.

The direct effect of levobupivacaine in isolated rat aorta involves lipoxygenase pathway activation and endothelial nitric oxide release.

BACKGROUND: Levobupivacaine is a long-acting local anesthetic with a clinical profile similar to that of racemic bupivacaine but with a greater margin of safety. Levobupivacaine produces dose-dependent vasoconstriction in vivo. Our goal in this in vitro study was to investigate the role of pathways involved in arachidonic acid metabolism in the levobupivacaine-induced contraction of isolated rat aorta and to determine which endothelium-derived vasodilators are involved in the modulation of levobupivacaine-induced contraction. METHODS: Rat thoracic aortic rings were isolated and suspended for isometric tension recording. Cumulative levobupivacaine dose-response curves over a range of 10(-6) to 3 x 10(-4) M were constructed in 1) aortic rings with no drug pretreatment; 2) endothelium-denuded rings pretreated with quinacrine dihydrochloride (nonspecific phospholipase A(2) inhibitor: 2 x 10(-5), 4 x 10(-5) M), nordihydroguaiaretic acid (NDGA) (lipoxygenase inhibitor: 10(-5), 3 x 10(-5) M), indomethacin (nonspecific cyclooxygenase inhibitor: 10(-5) M), AA-861 (5-lipoxygenase inhibitor: 10(-5), 5 x 10(-5) M), fluconazole (cytochrome P450 epoxygenase inhibitor: 10(-5) M), verapamil (10(-5) M), or calcium-free solution; and 3) endothelium-intact rings pretreated with N(omega)-nitro-L-arginine methyl ester (L-NAME) (nitric oxide synthase inhibitor: 5 x 10(-5) M), indomethacin, or fluconazole. Levobupivacaine-induced contractile response at each concentration (10(-4), 3 x 10(-4) M) was assessed in endothelium-denuded rings. Dose-response curves for potassium chloride in endothelium-denuded rings were generated in the presence or absence of NDGA and AA-861. Intracellular Ca(2+) levels were monitored by Ca(2+) image analysis using Fluo-4 fluorescence in vascular smooth muscle cells treated with levobupivacaine alone or AA-861 plus levobupivacaine. RESULTS: Levobupivacaine produced a tonic contraction in isolated rat aorta rings; this response was maximal at 10(-4) M levobupivacaine and gradually attenuated at 3 x 10(-4) M levobupivacaine. Levobupivacaine-induced contractions of endothelium-denuded rings were larger than those of endothelium-intact rings. Levobupivacaine-induced contraction of endothelium-denuded rings was attenuated by quinacrine dihydrochloride, NDGA, AA-861, verapamil, and calcium-free solution and, to a lesser extent, by indomethacin. L-NAME enhanced levobupivacaine-induced contraction of endothelium-intact rings and indomethacin slightly attenuated this contraction. NDGA and AA-861 attenuated the potassium chloride-induced contraction. AA-861 attenuated the levobupivacaine-induced intracellular calcium increase in vascular smooth muscle cells. CONCLUSIONS: Our data indicate that levobupivacaine-induced contraction of rat aortic smooth muscle is mediated mainly by activation of the lipoxygenase pathway and in part by activation of the cyclooxygenase pathway. In addition, activation of the lipoxygenase pathway seems to facilitate calcium influx via L-type calcium channels. Endothelial nitric oxide attenuates levobupivacaine-induced contraction.

A three-dimensional hyaluronic acid-based niche enhances the therapeutic efficacy of human natural killer cell-based cancer immunotherapy.

Adoptive transfer of natural killer (NK) cells is becoming one of the most important parts of cancer immunotherapy. However, recent accomplishments have focused on the improvement of the targeting effects based on the engineering of chimeric antigen receptors (CARs) on cell surfaces. Despite the large quantity of therapeutic cells required for clinical applications, the technology for ex vivo expansion is not well developed. Herein, a three-dimensional (3D) engineered hyaluronic acid-based niche for cell expansion (3D-ENHANCE) is introduced. Compared with the conventional two-dimensional (2D) method, NK-92 cell lines and human EGFR-specific (CAR)-NK cells cultured in 3D-ENHANCE yield favorable mRNA expressions, elevated cytokine release, upregulated proliferative and tumor-lytic abilities, and result in enhanced antitumor efficacy. Furthermore, controllable degradation rates can be realized by tuning the formulation of 3D-ENHANCE so that it can be applied as an implantable cell reservoir at surgical sites. In vivo results with the incompletely resected MDA-MB-231 model confirm that the peri-operative implantation of 3D-ENHANCE prevents the relapse and metastases after surgery. Overall, 3D-ENHANCE presents an effective cytokine-free niche for ex vivo expansion and postsurgical treatment that enhances the low-therapeutic efficacy of human NK cells.

Propofol limits rat myocardial ischemia and reperfusion injury with an associated reduction in apoptotic cell death in vivo.

Propofol, a rapidly acting, short duration, intravenous hypnotic anesthetic induction agent, is often used in clinical situations where myocardial ischemia/ reperfusion (I/R) injury is a threat. The aim of the present study was to evaluate the protective effect of propofol on myocardial I/R injury in rat due to apoptosis. Myocardial I/R injury were induced by occluding the left anterior descending (LAD) coronary artery for 25 min followed by either 2 h or 6 h reperfusion. Apoptosis was evaluated by Western blot analysis (Bcl-2, Bax expression), DNA strand breaks, TUNEL analysis and measuring myocardial caspase-3 activity. Propofol significantly reduced infarct size and improved I/R-induced myocardial contractile dysfunction by improving left ventricular diastolic pressure and positive and negative maximal values of the first derivative (+dp/dt) of left ventricular pressure. Propofol increased Bcl-2/Bax expression ratio and decreased caspase-3 activity in I/R rat hearts, which resulted in reduction of myocardial apoptosis as evidenced by TUNEL analysis and DNA laddering experiments. In an in vitro study, propofol increased H9c2 cell viability against oxidative stress induced by glucose oxidase (GOX) in a dose-dependent manner. These data suggest propofol limits I/R injury with an associated reduction in apoptotic cell death in vivo.

Nanoengineered Immune Niches for Reprogramming the Immunosuppressive Tumor Microenvironment and Enhancing Cancer Immunotherapy.

Cancer immunotherapies that harness the body's immune system to combat tumors have received extensive attention and become mainstream strategies for treating cancer. Despite promising results, some problems remain, such as the limited patient response rate and the emergence of severe immune-related adverse effects. For most patients, the therapeutic efficacy of cancer immunotherapy is mainly limited by the immunosuppressive tumor microenvironment (TME). To overcome such obstacles in the TME, the immunomodulation of immunosuppressive factors and therapeutic immune cells (e.g., T cells and antigen-presenting cells) should be carefully designed and evaluated. Nanoengineered synthetic immune niches have emerged as highly customizable platforms with a potent capability for reprogramming the immunosuppressive TME. Here, recent developments in nano-biomaterials that are rationally designed to modulate the immunosuppressive TME in a spatiotemporal manner for enhanced cancer immunotherapy which are rationally designed to modulate the immunosuppressive TME in a spatiotemporal manner for enhanced cancer immunotherapy are highlighted.

Lyophilizable and Multifaceted Toll-like Receptor 7/8 Agonist-Loaded Nanoemulsion for the Reprogramming of Tumor Microenvironments and Enhanced Cancer Immunotherapy.

The low therapeutic efficacy of current cancer immunotherapy is related to nonimmunogenic and immunosuppressive tumor microenvironments (TMEs). To overcome these limitations, both the immune priming of antitumoral lymphocytes and the reprogramming of immunosuppressive factors in TMEs are essential. Here, we suggest a nanoemulsion (NE)-based immunotherapeutic platform that can not only modulate tumor-induced suppression but also induce an effective cell-mediated immune response for T cell proliferation. Multifunctional NEs can be fabricated by integrating the efficacy of NEs as delivery systems and the multifaceted immunomodulation characteristics (i.e., immunostimulation and reprogramming of immunosuppression) of small molecule-based Toll-like receptor 7/8 agonists. Local in situ vaccination of melanoma and cervical tumor models with tumor antigens (protein and peptide) adjuvanted with NE loaded with TLR7/8 agonists [NE (TLR7/8a)] induced the recruitment and activation of innate immune cells, infiltration of lymphocytes, and polarization of tumor-associated M2 macrophages, which resulted in inhibition of tumor growth and prolonged survival in both primary and rechallenged tumor models. Antibody-depletion experiments also suggested that macrophages, type I IFN (IFN-α and IFN-ß), CD8+ T cells, and NK1.1+ cells contributed to the antitumor effect of NE (TLR7/8a). The combination of antitumoral lymphocytes and reprogramming of immunosuppressive TMEs induced by NE (TLR7/8a) treatment evoked a synergistic antitumor immune response with immune checkpoint blockade therapy (anti-PD-1 and anti-PD-L1).

Syringeable immunotherapeutic nanogel reshapes tumor microenvironment and prevents tumor metastasis and recurrence.

The low response rate of current cancer immunotherapy suggests the presence of few antigen-specific T cells and a high number of immunosuppressive factors in tumor microenvironment (TME). Here, we develop a syringeable immunomodulatory multidomain nanogel (iGel) that overcomes the limitation by reprogramming of the pro-tumoral TME to antitumoral immune niches. Local and extended release of immunomodulatory drugs from iGel deplete immunosuppressive cells, while inducing immunogenic cell death and increased immunogenicity. When iGel is applied as a local postsurgical treatment, both systemic antitumor immunity and a memory T cell response are generated, and the recurrence and metastasis of tumors to lungs and other organs are significantly inhibited. Reshaping of the TME using iGel also reverts non-responding groups to checkpoint blockade therapies into responding groups. The iGel is expected as an immunotherapeutic platform that can reshape immunosuppressive TMEs and synergize cancer immunotherapy with checkpoint therapies, with minimized systemic toxicity.

Lipid Emulsion Inhibits Apoptosis Induced by a Toxic Dose of Verapamil via the Delta-Opioid Receptor in H9c2 Rat Cardiomyoblasts.

The goals of this study were to investigate the effects of lipid emulsion (LE) on apoptosis induced by a toxic dose of verapamil in H9c2 cells and to elucidate the associated cellular mechanism. The effects of LE alone and combined with an inhibitor on the decreases in cell counts and viability induced by verapamil and diltiazem were examined using the MTT assay. The effects of verapamil alone, combined LE and verapamil treatment, and combined inhibitor, LE and verapamil treatment on cleaved caspase-3, caspase-8 and Bax expression, were examined using Western blotting. The effects of verapamil alone and combined with LE on the number of TUNEL-positive H9c2 cells were also examined. LE attenuated the decreases in cell counts and viability induced by verapamil and diltiazem. However, the magnitude of the LE-mediated attenuation of decreased cell viability was enhanced by verapamil compared with diltiazem treatment. Naloxone, naltrindole hydrochloride, LY294002 and MK-2206 inhibited the LE-mediated attenuation of increased cleaved caspase-3 and caspase-8 expression induced by verapamil. LE attenuated the increase in the number of TUNEL-positive cell induced by verapamil. These results suggest that LE attenuates apoptosis induced by verapamil via activation of the delta-opioid receptor, phosphoinositide 3-kinase and Akt.

Anti-apoptotic and myocardial protective effects of ethyl pyruvate after regional ischaemia/reperfusion myocardial damage in an in vivo rat model.

INTRODUCTION: The integration of reactive oxygen species is strongly associated with important pathophysiological mechanisms that mediate myocardial ischaemia/reperfusion (I/R) damage. Pyruvate is an efficacious scavenger of reactive oxygen species and a previous study has shown that ethyl pyruvate (EP) has a myocardial protective effect against regional I/R damage in an in vivo rat model. The purpose of this study was to determine whether the myocardial protective effect of EP is associated with anti-apoptosis. METHODS: Rats were allocated to receive EP dissolved in lactated Ringer's solution or lactated Ringer's solution alone, via intraperitoneal infusion one hour before ischaemia. They were exposed to 30 minutes of ischaemia followed by reperfusion of the left coronary artery territory over two hours. Anti-apoptotic effects were checked using several biochemical parameters after two hours of reperfusion. Apoptosis was analysed using measured caspase-3 activity, Western blotting of B-cell lymphoma 2 (Bcl-2) family protein cleaved by caspase-3, and assessment of DNA laddering patterns and the terminal deoxynucleotidyl transferase dUTP nick end labelling (TUNEL) staining test. RESULTS: In ischaemic myocardium, EP increased Bcl-2 expression, but reduced Bcl-2-associated X protein and cleaved caspase-3 expressions. EP reduced the expression of DNA laddering and the number of myocardial I/R-damaged TUNEL-positive cells. CONCLUSION: This study demonstrated that EP has an anti-apoptotic effect after regional I/R damage in an in vivo rat heart model. The myocardial protective effect of EP may be related to its anti-apoptotic effect.