The membrane depolarization and increase intracellular calcium level produced by silver nanoclusters are responsible for bacterial death

This work highlights how our silver ultra nanoclusters (ARGIRIUM-SUNc) hand-made synthesized, are very useful as a bactericide and anti-biofilm agent. The Argirium-SUNc effective antibacterial concentrations are very low (< 1 ppm) as compared to the corresponding values reported in the literature. Different bacterial defense mechanisms are observed dependent on ARGIRIUM-SUNc concentrations. Biochemical investigations (volatilome) have been performed to understand the pathways involved in cell death. By using fluorescence techniques and cell viability measurements we show, for the first time, that membrane depolarization and calcium intracellular level are both primary events in bacteria death. The ARGIRIUM-SUNc determined eradication of different biofilm at a concentration as low as 0.6 ppm. This suggests that the effect of the nanoparticles follows a common mechanism in different bacteria. It is highly probable that the chemical constitution of the crosslinks could be a key target in the disrupting mechanism of our nanoparticles. Since the biofilms and their constituents are essential for bacterial survival in contact with humans, the silver nanoparticles represent a logical target for new antibacterial treatments.

Modeling the spatiotemporal intracellular calcium dynamics in nerve cell with strong memory effects

Calcium signaling in nerve cells is a crucial activity for the human brain to execute a diversity of its functions. An alteration in the signaling process leads to cell death. To date, several attempts registered to study the calcium distribution in nerve cells like neurons, astrocytes, etc. in the form of the integer-order model. In this paper, a fractional-order mathematical model to study the spatiotemporal profile of calcium in nerve cells is assembled and analyzed. The proposed model is solved by the finite element method for space derivative and finite difference method for time derivative. The classical case of the calcium dynamics model is recovered by setting the fractional parameter and that validates the model for classical sense. The numerical computations have systematically presented the impact of a fractional parameter on nerve cells. It is observed that calbindin-D28k provides a significant effect on the spatiotemporal variation of calcium profile due to the amalgamation of the memory of nerve cells. The presence of excess amounts of calbindin-D28k controls the intracellular calcium level and prevents the nerve cell from toxicity.

Cav3.3 Down-Regulates CaMKII Beta to Ameliorate Neurotoxic Injury to the DRG Induced by Ropivacaine Hydrochloride

Background: The neurotoxicity of local anaesthetics is often reported. The present study aimed to investigate the relationship between Cav3.3 and CaMKⅡbeta in local anaesthetic neurotoxicity. Methods: An in vitro ropivacaine-induced model of neurotoxic injury to the dorsal root ganglion neurons was used. After specifically inhibiting the expression of Cav3.3 mRNA in DRG neurons by RNAi, cell viability, CaMKⅡbeta expression and the intracellular calcium ion level were detected in DRG neurons.Results: The results showed that the expression of CaMKⅡbeta decreased after the expression of Cav3.3 was inhibited. At the same time, in a model of ropivacaine hydrochloride-induced neurotoxic injury, inhibiting the expression of Cav3.3 mRNA improved cell viability and reduced the intracellular calcium level.Conclusions: These results suggest that Cav3.3 may be involved in neurotoxic injury induced by local anaesthetics by regulating the expression of CaMKⅡbeta. Both Cav3.3 and CaMKⅡbeta are therapeutic targets for neuronal injury induced by local anaesthetics.

Withdrawn: Safranal induces autophagy by AMPK activation and protects neurons against amyloid beta in Alzheimer’s disease

This article has been withdrawn by the Editor Purpose: To investigate autophagic induction by safranal and neuroprotection against amyloid beta in Alzheimer’s disease. Methods: Primary neurons and SH-SY5Y cells were used in this study. Assessment of cell proliferation and neuroprotection by safranal against amyloid beta was done by 3-(4,5-dimethylthiazol-2-yl)-2,5- diphenyltetrazolium bromide (MTT) assay. AMPK activation and mTOR inhibition were determined by western blot. Changes in intracellular calcium level, reactive oxygen species (ROS) and mitochondrial membrane potential (MMP) were assessed by flow cytometry. Results: Safranal protected neurons against amyloid beta toxicity. Furthermore, safranal activated AMPK pathway by activation of calcium/calmodulin-dependent protein kinase (CaMKKβ) to induce autophagy in both cell lines. The toxicity induced by amyloid beta in primary neurons and SH-SY5Y cells were attenuated by safranal. Moreover, amyloid beta-induced calcium levels were significantly decreased by safranal while ROS and MMP loss produced by amyloid beta was attenuated by safranal. Conclusion: These findings suggest that safranal protects neurons against amyloid beta by inducing autophagy via AMPK pathway. Therefore, safranal is a probable therapeutic target for Alzheimer’s disease.

Derrone Inhibits Platelet Aggregation, Granule Secretion, Thromboxane A2 Generation, and Clot Retraction: An In Vitro Study

Cudrania tricuspidata (C. tricuspidata) is widespread throughout East Asia and in China and Korea, and it is widely used as a traditional remedy against eczema, mumps, and tuberculosis. With regard to the aforementioned medical efficacy, various studies are continuously being conducted, and it has been reported that C. tricuspidata extract has various actions against inflammation, diabetes, obesity, and tumors. Therefore, we evaluated antiplatelet effects using derrone in C. tricuspidata. We examined the effect of derrone on the phosphorylation of vasodilator-stimulated phosphoprotein (VASP) and inositol 1, 4, 5-triphosphate receptor I (IP3RI), and on the dephosphorylation of cytosolic phospholipase A2 (cPLA2), mitogen-activated protein kinases p38 (p38MAPK), and Akt, which affects platelet function and thrombus formation. Various agonists-induced human platelets were inhibited by derrone without cytotoxicity, and it also decreased the intracellular calcium level through the signaling molecule phosphorylations. In addition, derrone inhibited glycoprotein IIb/IIIa (αIIb/β3) affinity. Thus, in the present study, derrone suppressed human platelet aggregation and thrombin-induced clot formation.

Secondary Metabolites of Plants as Modulators of Endothelium Functions

According to the World Health Organization, cardiovascular diseases are the main cause of death worldwide. They may be caused by various factors or combinations of factors. Frequently, endothelial dysfunction is involved in either development of the disorder or results from it. On the other hand, the endothelium may be disordered for other reasons, e.g., due to infection, such as COVID-19. The understanding of the role and significance of the endothelium in the body has changed significantly over time—from a simple physical barrier to a complex system encompassing local and systemic regulation of numerous processes in the body. Endothelium disorders may arise from impairment of one or more signaling pathways affecting dilator or constrictor activity, including nitric oxide–cyclic guanosine monophosphate activation, prostacyclin–cyclic adenosine monophosphate activation, phosphodiesterase inhibition, and potassium channel activation or intracellular calcium level inhibition. In this review, plants are summarized as sources of biologically active substances affecting the endothelium. This paper compares individual substances and mechanisms that are known to affect the endothelium, and which subsequently may cause the development of cardiovascular disorders.

CRISPR/Cas9-mediated Tryptophan Hydroxylase 1 Knockout Decreases Calcium Transportation in Goat Mammary Epithelial Cells

Background: Calcium is one of the major mineral nutrients in goat milk. Tryptophan hydroxylase1 (TPH1) is a rate-limiting enzyme catalyzing hydroxylation of L-tryptophan into 5-hydroxytryptamine (5-HT) essential for maintaining calcium homeostasis. The function of TPH1 and 5-HT in goat mammary calcium homeostasis is not well known.Methods: The CRISPR/Cas9-meidated TPH1 knockout goat mammary epithelial cells (GMEC) were constructed firstly. Then the content of 5-HT, intracellular calcium level and abundance of key genes related to calcium transportation were evaluated and compared in wild-type GMEC, TPH1 knockout GMEC, to explore the impact of TPH1 on calcium transportation, respectively. Wild-type GMEC and TPH1 knockout GMEC were further treated with exogenous 5-HTP to confirm the role of TPH1 in regulating calcium homeostasis in GMEC. 5-HT concentration was measured by enzyme-linked immunosorbent assay and fluo-3 staining was used to determine intracellular calcium content.Results: The TPH1 knockout GMEC heterozygous clone with no off-target effects was obtained after transfection of the Cas9/sgRNA expression vector. The 5-HT synthesis and intracellular calcium level decreased in TPH1 gene knockout GMEC. The mRNA abundance of secretory-pathway Ca2+ -ATPase1 (SPCA1) and plasma membrane Ca2+-ATPase1 (PMCA1) were up-regulated while the mRNA abundance of secretory-pathway Ca2+ -ATPase2 (SPCA2) was down-regulated in TPH1 knockout GMEC. Up-regulation of parathyroid hormone-related peptide (PTHrP), a key regulator of mammary calcium metabolism, induced by 5-HTP were blocked by TPH1 gene knockout. The TPH1 knockout GMEC showed a lower sensitivity to 5-HTP induced elevation of calcium content.Conclusion: Results suggested that TPH1 plays an important role in regulating calcium homeostasis via PTHrP and calcium transportation related factors in GMEC.

Abstract 15: The Role Of Opioid Receptors In Podocytes In The Development Of Hypertension In Dahl Salt-sensitive Rats

The rise in opioid use underscores the importance to better understand the direct and indirect effects of opioids on renal function and blood pressure. Although opioid use is associated with predictors of cardiovascular diseases, these drugs are common analgesics for hypertensive patients. We hypothesize that stimulation of opioid receptors (ORs) leads to elevated intracellular calcium level in podocytes ultimately leading to cell apoptosis, development of albuminuria and consequent progression of hypertension. Live calcium imaging experiments on freshly isolated glomeruli from rat and human kidneys, as well as human immortalized podocyte cell line, was performed to test the effect of specific ORs agonists. Following experiments assessed the effect of opioid signaling on the development of hypertension and kidney function in Dahl salt-sensitive (SS) rats, which were fed a 0.4% (LS) or 8% (HS) NaCl diets for 14 days with or without a daily i.v. bolus infusion of BRL52537, a potent and selective kappa-OR agonist. Stimulation of kappa-ORs, but not mu-ORs or delta-ORs, mediated calcium influx in podocytes through activation of TRPC6 channels. The effect of BRL52537 was completely abolished when we used the 0 mM calcium media or when a TRPC6 channel inhibitor (SAR7334) was applied. Triggering the kappa-OR/TRPC6 pathway induced podocyte cell shape changes via actin cytoskeleton remodeling. In vivo studies revealed that rats chronically treated with BRL52537 exhibited augmented blood pressure (MAP was 179 ± 15 vs. 151 ± 11 mmHg), albuminuria, and elevation in podocyte calcium. Western blot analysis revealed elevated levels of nephrin in urine samples and pro-caspase-3 in renal cortex. Moreover, TRPC6 expression was elevated under hypertensive conditions and further promoted pathological increase in calcium influx in response to kappa-OR stimulation. Summarizing the data, the opioid-induced increase in the calcium flux in podocytes is expected to contribute to kidney injury leading to progression of salt-induced hypertension. These data demonstrate that the kappa-OR/TRPC6 signaling pathway directly influences podocyte calcium handling, provoking the development of kidney injury in the opioid treated hypertensive cohort.

Calcium spikes accompany the cleavage furrow ingression and cell separation during fission yeast cytokinesis

The role of calcium signaling during cytokinesis has long remained ambiguous. The studies of embryonic cell division discovered that calcium concentration increases transiently at the division plane just before the cleavage furrow ingression, leading to the hypothesis that these calcium transients trigger the contractile ring constriction. However, such calcium transients have only been found in animal embryos and their function remains controversial. Here we explored cytokinetic calcium transients in the model organism fission yeast. We adopted GCaMP, a genetically encoded calcium indicator, to determine the intracellular calcium level. We validated GCaMP as a highly sensitive calcium indicator which allowed us to capture the calcium transients stimulated by osmotic shocks. To identify calcium transients during cytokinesis, we first identified a correlation between the intracellular calcium level and cell division. Next, we discovered calcium spikes at the start of the cleavage furrow ingression and the end of the cell separation using time-lapse microscopy to. Inhibition of these calcium spikes slowed down the furrow ingression and led to frequent lysis of the daughter cells. We conclude that like the larger animal embryos fission yeast triggers cytokinetic calcium transients which promote the ring constriction and daughter cell integrity (194).Highlight summary for TOCCalcium rises transiently at the division plane during embryonic cell cytokinesis, but the conservation and function of such calcium transients remain unclear. We identified similar calcium spikes during fission yeast cytokinesis and demonstrated that these spikes promote the contractile ring constriction and the daughter cell integrity (257).