The effects of immune cells, in particular macrophages, on the behaviour of mesenchymal stromal cells (MSCs) have recently gained much attention for MSCs-based tissue-engineered constructs. This study aimed to evaluate the effect of monocytes/macrophages on the osteogenic differentiation of adipose-derived mesenchymal stromal cells (ADMSCs) in three-dimensional (3D) cocultures. For this, we cocultured THP-1 monocytes, M1 macrophages, or M2 macrophages with ADMSCs on 3D poly(lactic-co-glycolic) acid (PLGA)/polycaprolactone (PCL) scaffolds using osteogenic medium for up to 42 days. We found that osteogenic differentiation of ADMSCs was inhibited by monocytes and both macrophage subtypes in 3D scaffolds. Furthermore, coculture of monocytes/macrophages with ADMSCs resulted in downregulated secretion of oncostatin M (OSM) and bone morphogenetic protein 2 (BMP-2) and inhibited expression of osteogenic markers alkaline phosphatase (ALP), bone sialoprotein (BSP), and runt-related transcription factor 2 (RUNX2). Compared with both macrophage subtypes, monocytes inhibited osteogenic differentiation of ADMSCs more significantly. These data suggest that the mutual interactions between monocytes/macrophages and ADMSCs negatively affect MSC osteogenic differentiation and thus possibly bone healing capacity, which highlights the importance of the micro-environment in influencing cell-based constructs to treat bone defects and the potential to improve their performance by resolving the inflammation ahead of treatment.

The embryonic origin of pericytes is heterogeneous, both between and within organs. While pericytes of coelomic organs were proposed to differentiate from the mesothelium, a single-layer squamous epithelium, the embryonic origin of pancreatic pericytes has yet to be reported. Here, we show that adult pancreatic pericytes originate from the embryonic pancreatic mesenchyme. Our analysis indicates that pericytes of the adult mouse pancreas originate from cells expressing the transcription factor Nkx3.2. In the embryonic pancreas, Nkx3.2-expressing cells constitute the multilayered mesenchyme, which surrounds the pancreatic epithelium and supports multiple events in its development. Thus, we traced the fate of the pancreatic mesenchyme. Our analysis reveals that pancreatic mesenchymal cells acquire various pericyte characteristics, including gene expression, typical morphology, and periendothelial location, during embryogenesis. Importantly, we show that the vast majority of pancreatic mesenchymal cells differentiate into pericytes already at embryonic day 13.5 and progressively acquires a more mature pericyte phenotype during later stages of pancreas organogenesis. Thus, our study indicates the embryonic pancreatic mesenchyme as the primary origin to adult pancreatic pericytes. As pericytes of other coelomic organs were suggested to differentiate from the mesothelium, our findings point to a distinct origin of these cells in the pancreas. Thus, our study proposes a complex ontogeny of pericytes of coelomic organs.

Reactive oxygen species (ROS) play a key role in development of heart failure but, at a cellular level, their effects range from cytoprotection to induction of cell death. Understanding how this is regulated is crucial to develop novel strategies to ameliorate only the detrimental effects. Here, we revisited the fundamental hypothesis that the level of ROS per se is a key factor in the cellular response by applying different concentrations of H2O2 to cardiomyocytes. High concentrations rapidly reduced intracellular ATP and inhibited protein synthesis. This was associated with activation of AMPK which phosphorylated and inhibited Raptor, a crucial component of mTOR complex-1 that regulates protein synthesis. Inhibition of protein synthesis by high concentrations of H2O2 prevents synthesis of immediate early gene products required for downstream gene expression, and such mRNAs (many encoding proteins required to deal with oxidant stress) were only induced by lower concentrations. Lower concentrations of H2O2 promoted mTOR phosphorylation, associated with differential recruitment of some mRNAs to the polysomes for translation. Some of the upregulated genes induced by low H2O2 levels are cytoprotective. We identified p21Cip1/WAF1 as one such protein, and preventing its upregulation enhanced the rate of cardiomyocyte apoptosis. The data support the concept of a "redox rheostat" in which different degrees of ROS influence cell energetics and intracellular signalling pathways to regulate mRNA and protein expression. This sliding scale determines cell fate, modulating survival vs death.

Aegle marmelos (L.) Correa (A. marmelos) has been used in Ayurvedic medicine as a brain tonic however its neuroprotective effect against streptozotocin (STZ) induced cognitive impairment and oxidative stress has not been reported yet in vivo. Therefore, the present study was attempted to investigate the neuroprotective potential of ethanolic extract of A. marmelos leaves (AME) on STZ induced memory impairment in male rats.

Evidence is accumulating that activation of mineralocorticoid (MR) and glucocorticoid (GR) receptors on peripheral sensory neurons modulates pain sensation. While the expression and exact anatomical localization of MR and GR in the various subpopulations of peripheral sensory neurons has been shown in animals, it is still unknown for the human skin. Therefore, we aimed to identify MR and GR mRNA and protein as well as the exact subpopulations of sensory neurons in human versus rat skin. Tissue samples from rat and human skin were subjected to RT-PCR, Western blot and double immunofluorescence confocal analysis of MR and GR with the neuronal markers calcitonin gene-related peptide (CGRP), neurofilament 200 (NF200) and tyrosine hydroxylase (TH). Using RT-PCR we were able to isolate MR as well as GR specific transcripts from human skin. Consistently, Western blot analysis identified MR- as well as GR- specific protein bands at the expected molecular weights of 110 and 87 kD, respectively. Double immunofluorescence confocal microscopy of human skin revealed that MR predominantly colocalized with calcitonin-gene-related peptide (CGRP)-immunoreactive (IR) nociceptive neurons - similar to rat skin - underscoring a pivotal role for MR in the modulation of pain. The majority of GR-immmunoreactivity was localized in peripheral peptidergic CGRP-IR sensory nerve fibers, but in addition on TH-IR sympathetic postganglionic, and NF200-IR myelinated mechanoreceptive nerve fibers, both within human and rat skin. Moreover, GR but not MR were localized in keratinocytes of the epidermal layer of human and rat skin. Overall, our results indicate considerable overlap in sensory neuron expression of MR and GR in humans and rats endorsing a common systems approach in mammals that may modulate the transmission of sensory information by MR and GR activation.

The NLRP3 inflammasome is activated in the cytoplasm of cells and its products such as IL-1β are exported through a non-classical ER-Golgi pathway. Several mechanistically distinct models including exocytosis of secretory lysosomes, microvesicles (MVs) and extracellular vehicles (EVs) have been proposed for their release. In this study, we hypothesized that the NLRP3 inflammasome product, IL-1β in response to exogenously administrated and endogenously produced d-ribose stimulation is released via extracellular vesicles including EVs via a sphingolipid-mediated molecular mechanisms controlling lysosome and multivesicular body (MVB) interaction. First, we demonstrated that both endogenous and exogenous d-ribose induced NLRP3 inflammasome activation to produce IL-1β, which was released via EVs in podocytes. Then, we found that colocalization of marker MVB marker VPS16 with IL-1β within podocytes increased upon d-ribose stimulation, which was accompanied by decreased colocalization of lysosome marker Lamp-1 and VPS16, suggesting decrease in MVB inclusion of IL-1β due to reduced lysosome and MVB interaction. All these changes were mimicked and accelerated by lysosome v-ATPase inhibitor, bafilomycin. Moreover, ceramide in podocytes was found elevated upon d-ribose stimulation, and prior treatments of podocyte with acid sphingomyelinase (Asm) inhibitor, amitriptyline, acid ceramidase (AC) inducer, genistein, or AC CRISPR/cas9 activation plasmids were found to decrease d-ribose-induced ceramide accumulation, EVs release and IL-1β secretion due to reduced interactions of lysosome with MVBs. These results suggest that inflammasome-derived products such as IL-1β during d-ribose stimulation are released via EVs, in which lysosomal sphingolipid-mediated regulation of lysosome function plays an important role.

Hearing loss and cognitive decline are commonly associated with aging and morbidity. Present clinical interest lies in whether peripheral hearing loss promotes cognitive decline and if prophylaxis with selective adenosine receptor agonist CGS21680 effectively mitigates the adverse effects. In the current study, male Sprague Dawley rats weighing 200-250 g m were randomly allocated into three groups: Group 1) rats exposed to 100 dB SPL white noise, 2 h a day for 15 consecutive days, 2) rats supplemented with an adenosine receptor agonist, CGS21680 at 100 μg/kg/day prior to noise exposure and 3) unexposed control rats. Baseline hearing and cognition assessed by auditory brainstem response (ABR) and water maze respectively was undertaken for all the groups. Phalloidin stain and synaptic ribbons count in cochlea, and, Ki67, DCX and NeuN in hippocampus were observed by immunohistochemistry. It was inferred that noise exposed rats showed elevated thresholds of ABR and poorer performances in spatial working memory when compared with controls. On the contrary, CGS21680 administered group exhibited improved ABR and cognitive functions with shorter mean latency and path-length to reach the platform, significant reduction in the noise induced loss of synaptic ribbons count and increased number of Ki67 and doublecortin (DCX) positive cells compared to their noise exposed counterparts. Pharmacologic intervention with selective A2A receptor agonist CGS21680 provided adequate protection from noise by effectively maintaining hearing threshold levels, cell viability in cochlea and hippocampus & functional/intact reference memory.

Gene-silencing with targeted siRNAs has great potential as a therapeutic approach for various diseases including cancer. However, intracellular delivery of siRNA is challenging. We used bovine milk exosomes as a novel system for siRNA delivery. First, we demonstrated that exosomes can deliver endogenous RNA payloads into recipient cells. Next, we loaded siRNA against specific genes including VEGF, EGFR, AKT, MAPK, and KRAS. We utilized 5'-32P-labeled siKRAS as a tracer and found exosome loading with siRNA could be variable. We demonstrated that the siRNA of loaded exosomes is stable and resist degradation. Our results indicated that siRNAs against target genes ranged from 2 to 10-fold knockdown in expression levels in various cancers. Since mutated KRAS has been implicated in the development of various cancers including lung cancer, we tested a mutant-allele specific siRNA against KRASG12S, in A549 cells. We observed a dose-dependent anti-proliferative activity against A549 cells treated with exosomes carrying siKRASG12S. We observed significant inhibition of A549 tumor xenografts in animals treated with folic acid-functionalized exosomes carrying siKRAS. In summary, milk-derived exosomes represent a viable natural nano-carrier for the delivery of siRNA for therapeutic application against cancer.

Wnt signaling is overactivated in triple-negative breast cancer (TNBC) and several other cancers, and its suppression emerges as an effective anticancer treatment. However, no drugs targeting the Wnt pathway exist on the market nor in advanced clinical trials. Here we provide a comprehensive body of preclinical evidence that an anti-leprotic drug clofazimine is effective against TNBC. Clofazimine specifically inhibits canonical Wnt signaling in a panel of TNBC cells in vitro. In several mouse xenograft models of TNBC, clofazimine efficiently suppresses tumor growth, correlating with in vivo inhibition of the Wnt pathway in the tumors. Clofazimine is well compatible with doxorubicin, exerting additive effects on tumor growth suppression, producing no adverse effects. Its excellent and well-characterized pharmacokinetics profile, lack of serious adverse effects at moderate (yet therapeutically effective) doses, its combinability with cytotoxic therapeutics, and the novel mechanistic mode of action make clofazimine a prime candidate for the repositioning clinical trials. Our work may bring forward the anti-Wnt targeted therapy, desperately needed for thousands of patients currently lacking targeted treatments.

Tumor-initiating cells (TICs), which are responsible for sustaining tumor growth and recurrence, rely on several regulatory factors. However, the mechanism of inflammation-related molecules in the acquisition and maintenance of TIC properties in hepatocellular carcinoma (HCC) remains elusive. We previously demonstrated that the voltage-gated calcium channel α2δ1 subunit is a functional surface marker of HCC TICs. Here, we found that the expression of an inflammation-related molecule C-X-C motif chemokine 11 (CXCL11) was significantly upregulated in α2δ1+ HCC TICs and that CXCL11 induced the expression of stem cell-related genes, such as BMI1, NANOG, MDR1, ABCG2, and CACNA2D1. Furthermore, CXCL11 could promote the acquisition and maintenance of self-renewal, tumorigenic, and chemoresistance properties of α2δ1+ HCC TICs by activating the extracellular signal-regulated kinase (ERK1/2) through its affinity receptor CXCR3. Collectively, our results suggest that CXCL11 may positively regulate the stemness of α2δ1+ HCC TICs via ERK1/2 activation through an autocrine signaling pathway.

In this review we will discuss how computational methods, and in particular classical molecular dynamics simulations, can be used to calculate solubility of pharmaceutically relevant molecules and systems. To the extent possible, we focus on the non-technical details of these calculations, and try to show also the added value of a more thorough and detailed understanding of the solubilization process obtained by using computational simulations. Although the main focus is on classical molecular dynamics simulations, we also provide the reader with some insights into other computational techniques, such as the COSMO-method, and also discuss Flory-Huggins theory and solubility parameters. We hope that this review will serve as a valuable starting point for any pharmaceutical researcher, who has not yet fully explored the possibilities offered by computational approaches to solubility calculations.

Opiate withdrawal induces an early aversive state which can be associated to contexts and/or cues, and re-exposure to either these contexts or cues may participate in craving and relapse. Nucleus accumbens (NAC), hippocampus (HPC) and basolateral amygdala (BLA) are crucial substrates for acute opiate withdrawal, and for withdrawal memory retrieval. Also HPC and BLA interacting with the NAC are suggested to respectively mediate the processing of context and cue representations of drug-related memories. Here we used a paradigm of conditioned suppression of operant food seeking, allowing to differentiate context and cue related responses, to study the influence of withdrawal memories on operant behavior and the underlying neural substrates. catFISH for Arc mRNA expression was used to discriminate cellular responses during context and cue (flashing light) periods in this paradigm. We show that reactivation of the memory of the negative affective state of withdrawal suppresses active lever pressing for food, and this conditioned suppression is generalized to the context. Interestingly the behavioral responses during the context and cue light periods are associated with differential Arc mRNA activations within the NAC, BLA, and HPC. Indeed both periods led to NAC shell activation whereas the NAC core was responsive only following the cue light period. Moreover, BLA and HPC were more responsive during cue-light and context period respectively. These data further support the already reported differential role of these brain structures on cue vs context-induced reinstatement of operant behaviors, and highlight the existence of common mechanisms for the processing of positive and aversive emotional memories.

Natural killer (NK) cells are innate immune lymphocytes with a key role in host defense against HIV infection. Recent advances in chimeric antigen receptors (CARs) have made NK cells a prime target for expressing recombinant receptors capable of redirecting NK cytotoxic functions towards HIV-infected cells. In this review, we discuss the role of NK cells in HIV and the mechanisms of actions of HIV-targeting CAR strategies. Furthermore, we also review NK cells signal transduction and its application to CAR NK cell strategies to develop new combinations of CAR intracellular domains and to improve CAR NK signaling and cytotoxic functions.

Several studies have documented the HPV genotypes in the Senegalese general population. The objective was to explore the HPV genotype distribution in Senegalese FSWs in order to assess the potential relevance of currently-available vaccines.

Domestic rice (Oryza sativa L.) is one of the most important cereal crops, feeding a large number of worldwide populations. Along with various high-throughput genome sequencing projects, rice genomics has been making great headway toward direct field applications of basic research advances in understanding the molecular mechanisms of agronomical traits and utilizing diverse germplasm resources. Here, we briefly review its achievements over the past two decades and present the potential for its bright future.

Ribes alpestre Decne has been commonly used in the treatment of joint complaints.

Chemotherapy of East Coast fever, a lymphoproliferative cancer-like disease of cattle causing significant economic losses in Africa, is largely dependent on the use of buparvaquone, a drug that was developed in the late 1980's. The disease is caused by the tick-borne protozoan pathogen Theileria parva. Buparvaquone can be used prophylactically and it is also active against tropical theileriosis, caused by the related parasite Theileria annulata. Recently, drug resistance was reported in T. annulata, and could occur in T. parva. Using a 3H-thymidine incorporation assay we screened 796 open source compounds from the Medicines for Malaria Venture (MMV) to discover novel chemicals with potential inhibitory activity to T. parva. We identified nine malaria box compounds and eight pathogen box compounds that inhibited the proliferation of F100TpM, a T. parva infected lymphocyte cell line. However, only two compounds, MMV008212 and MMV688372 represent promising leads with IC50 values of 0.78 and 0.61 μM, respectively, and CC50 values > 5 μM. The remaining compounds exhibited a high degree of toxicity (CC50 values < 1.09 μM) on the proliferation of bovine peripheral blood mononuclear cells stimulated with concanavalin A. We also tested the anti-cancer drug, dasatinib, used in the chemotherapy of some leukemias. Dasatinib was as active and safe as buparvaquone in vitro, with an IC50 of 5 and 4.2 nM, respectively, and CC50 > 10 μM. Our preliminary data suggest that it may be possible to repurpose compounds from the cancer field as well as MMV as novel anti-T. parva molecules.

The clinical efficiency of everolimus, an mammalian target of rapamycin (mTOR) inhibitor, is palliative as sequential or second-line therapy for renal cell carcinoma (RCC). However, the limited response of everolimus in RCC remains uncertain. In the present study, everolimus-resistant RCC models were established to understand the mechanisms and to seek combination approaches. Consequently, the activation of ERK was found to contribute toward everolimus-acquired resistance and poor prognosis in patients with RCC. In addition, the efficacy and mechanism of combination treatment underlying RCC using everolimus and ERK inhibitors was investigated. The ERK inhibitor in combination with everolimus synergistically inhibited the proliferation of RCC cells by arresting the cell cycle in the G1 phase. The combination treatment markedly attenuated the deoxyribonucleoside triphosphate (dNTP) pools by downregulating the mRNA expression of RRM1 and RRM2 through E2F1. The overexpression of E2F1 or supplementation of dNTP rescued the anti-proliferation activity of the everolimus-SCH772984 combination. The antitumor efficacy of combination therapy was reiterated in RCC xenograft models. Thus, the current findings provided evidence that the everolimus-ERK inhibitor combination is a preclinical therapeutic strategy for RCC.

Chemoresistance remains a great obstacle in effective lung adenocarcinoma (LUAD) treatment. Previously, we verified the role of microRNA-200b (miR-200b) in the formation of docetaxel (DTX)-resistant LUAD cells. This study aims to investigate the mechanism underlying the low level of miR-200b in DTX-resistant LUAD cells. The real-time reverse transcription (RT2) lncRNA PCR array system was applied to explore lncRNAs that potentially regulated miR-200b in DTX-resistant LUAD cells. Metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) contributed to the low miR-200b level in DTX-resistant LUAD cells. Functional assays were conducted to determine the role of MALAT1 in regulating the growth and metastasis of parental and DTX-resistant LUAD cells. Investigation revealed the mechanism of the competing endogenous RNA (ceRNA) pathway. MALAT1 regulated miR-200b by acting as a ceRNA. MALAT1 modulated the sensitivity of LUAD cells to DTX. E2F transcription factor 3 (E2F3) and zinc-finger E-box binding homeobox 1 (ZEB1) were two targets of miR-200b and mediated the function of MALAT1 in DTX-resistant LUAD cells. Transcription factor AP-2 gamma (TFAP2C) and ZEB1 activated the MALAT1 transcription. In conclusion, TFAP2C-activated MALAT1 modulated the chemoresistance of LUAD cells by sponging miR-200b to upregulate E2F3 and ZEB1. Our findings may provide novel therapeutic targets and perspectives for LUAD treatment.

GPR120 and GPR40 were recently reported as omega-3 (ω3) receptors with anti-inflammatory properties. Physical exercise could increase the expression of these receptors in the liver, improving hepatic metabolism in obesity and type 2 diabetes. Our aim was to investigate GPR120/40 in the liver of lean and obese mice after acute or chronic physical exercise, with or without the supplementation of ω3 rich flaxseed oil (FS), as well as assess the impact of exercise and FS on insulin signaling and inflammation. Mice were fed a high-fat diet (HF) for 4 weeks to induce obesity and subsequently subjected to exercise with or without FS, or FS alone. Insulin signaling, inflammatory markers and GPR120/40 and related cascades were measured. Chronic, but not acute, exercise and FS increased GPR120, but not GPR40, activating β-arrestin-2 and decreasing the inflammatory response, as well as reducing fat depots in liver and adipose tissue. Exercise or a source of ω3 led to a higher tolerance to fatigue and an increased running distance and speed. The combination of physical exercise and ω3 food sources could provide a new strategy against obesity through the modulation of hepatic GPR120 and an increase in exercise performance.