Neuropharmacological and Antidiarrheal Potentials of Duabanga grandiflora (DC.) Walp. Stem Bark and Prospective Ligand–Receptor Interactions of Its Bioactive Lead Molecules

Duabanga grandiflora (DC.) Walp. is an ethnomedicinally significant plant used to treat various illnesses, but there is little scientific evidence to support its use. This study explored the pharmacological activities of methanol extract of D. grandiflora stem barks (MEDG) through in vivo approaches in Swiss albino mice and a computer-aided molecular approach. The forced swimming test (FST), tail suspension test (TST), elevated plus maze (EPM), and hole board test (HBT) were used to determine anti-depressant and anxiolytic activity in experimental mice. In addition, anti-diarrheal studies were performed using castor oil-induced diarrhea, castor oil-induced enter pooling, and the charcoal-induced gastrointestinal motility test. MEDG showed substantial depletions in the immobility times in both FST and TST after treatment with the MEDG extract, whereas moderate anxiolytic activity was manifested at a higher dose (400 mg/kg) compared with the control. Correspondingly, MEDG extract revealed a significant reduction in wet feces and decreased the small intestinal transit of charcoal meal in castor oil-induced diarrhea and charcoal-induced gastrointestinal motility test. In the computer-aided molecular approaches, vanillin displayed a promising binding score for both anxiolytic and anti-diarrheal activities, while duabanganal C showed a promising score for the anti-depressant activity. The present experimental findings along with a computer-aided model conclude that MEDG could be a possible Phyto therapeutic agent with potential anti-depressant, anxiolytic and anti-diarrheal activity.     

A comparative study of carbonic anhydrase activity in lymphocytes from colorectal cancer tissues and adjacent healthy counterparts

Several carbonic anhydrase (CA, EC 4.2.1.1) isoforms play an essential role in processes connected to tumorigenesis, as they efficiently accelerate the hydration of carbon dioxide to bicarbonate and proton. In this context, examples are CA IX and CA XII, which were proved to be upregulated in many solid malignancies. On the other hand, cancer and the immune system are inextricably linked, and targeting the immune checkpoints recently was shown to efficiently improve the treatment of malignancies. In this study, we have investigated the expression of CA isoforms in tumour-infiltrating lymphocytes (TILs) that, according to the immunosurveillance theory, were suggested to have a crucial role in the development of colorectal cancer (CRC). T lymphocytes isolated from healthy surrounding mucosa showed a higher CA activity compared to those present in tumour and peripheral blood in the same patients. CA I and II were confirmed as enzyme isoforms involved in the process, as determined by proteomic analysis of corresponding TIL samples. These preliminary findings suggest a dysregulation of the local immune response in the CRC tissues and a loss of effective anticancer mechanisms mediated by CAs therein.     

Endocannabinoid dysregulation in the pancreas and adipose tissue of mice fed with a high-fat diet

Objective: In mice, endocannabinoids (ECs) modulate insulin release from pancreatic β‐cells and adipokine expression in adipocytes through cannabinoid receptors. Their pancreatic and adipose tissue levels are elevated during hyperglycemia and obesity, but the mechanisms underlying these alterations are not understood. Methods and Procedures: We assessed in mice fed for up to 14 weeks with a standard or high‐fat diet (HFD): (i) the expression of cannabinoid receptors and EC biosynthesizing enzymes (N‐acyl‐phosphatidyl‐ethanolamine‐selective phospholipase D (NAPE‐PLD) and DAGLα) and degrading enzymes (fatty acid amide hydrolase (FAAH) and monoacylglycerol lipase (MAGL)) in pancreatic and adipose tissue sections by immunohistochemical staining; (ii) the amounts, measured by liquid chromatography–mass spectrometry, of the ECs, 2‐AG, and anandamide (AEA). Results: Although CB₁ receptors and biosynthetic enzymes were found mostly in α‐cells, degrading enzymes were identified in β‐cells. Following HFD, staining for biosynthetic enzymes in β‐cells and lower staining for FAAH were observed together with an increase of EC pancreatic levels. While we observed no diet‐induced change in the intensity of the staining of EC metabolic enzymes in the mesenteric visceral fat, a decrease in EC concentrations was accompanied by lower and higher staining of biosynthesizing enzymes and FAAH, respectively, in the subcutaneous fat. No change in cannabinoid receptor staining was observed following HFD in any of the analyzed tissues. Discussion: We provide unprecedented information on the distribution of EC metabolic enzymes in the pancreas and adipose organ, where their aberrant expression during hyperglycemia and obesity contribute to dysregulated EC levels.     

Inhibition of tumor-associated human carbonic anhydrase isozymes IX and XII by a new class of substituted-phenylacetamido aromatic sulfonamides

Here, we investigate 28 structurally new sulfonamides and their subsequent testing for enzyme inhibition of cytosolic and tumor-associated carbonic anhydrases (CAs, EC 4.2.1.1). The compounds showed very potent inhibition of four physiologically relevant human (h) CA isoforms, namely hCA I, II, IX and XII. Interestingly, the K_I values were in the nanomolar range for the tumor-associated hCA IX and hCA XII. Docking studies have revealed details regarding the very favorable interactions between the scaffolds of this new class of inhibitors and the active sites of the investigated CA isoforms. As there are reported cases of tumors overexpressing both CA II and IX, such potent inhibitors for the two isoforms as those detected in this work, may have applications for targeting more than one CA present in tumors.     

Anti‐proliferative effect of rhein,an anthraquinone isolated from Cassia species,on Caco‐2 human adenocarcinoma cells

In recent years, the use of anthraquinone laxatives, in particular senna, has been associated with damage to the intestinal epithelial layer and an increased risk of developing colorectal cancer. In this study, we evaluated the cytotoxicity of rhein, the active metabolite of senna, on human colon adenocarcinoma cells (Caco‐2) and its effect on cell proliferation. Cytotoxicity studies were performed using 3‐(4,5‐dimethylthiazol‐2‐yl)‐2,5‐diphenyltetrazolium bromide (MTT), neutral red (NR) and trans‐epithelial electrical resistance (TEER) assays whereas ³H‐thymidine incorporation and Western blot analysis were used to evaluate the effect of rhein on cell proliferation. Moreover, for genoprotection studies Comet assay and oxidative biomarkers measurement (malondialdehyde and reactive oxygen species) were used. Rhein (0.1–10 μg/ml) had no significant cytotoxic effect on proliferating and differentiated Caco‐2 cells. Rhein (0.1 and 1 μg/ml) significantly reduced cell proliferation as well as mitogen‐activated protein (MAP) kinase activation; by contrast, at high concentration (10 μg/ml) rhein significantly increased cell proliferation and extracellular‐signal‐related kinase (ERK) phosphorylation. Moreover, rhein (0.1–10 μg/ml): (i) did not adversely affect the integrity of tight junctions and hence epithelial barrier function; (ii) did not induce DNA damage, rather it was able to reduce H₂O₂‐induced DNA damage and (iii) significantly inhibited the increase in malondialdehyde and reactive oxygen species (ROS) levels induced by H₂O₂/Fe²⁺. Rhein was devoid of cytotoxic and genotoxic effects in colon adenocarcinoma cells. Moreover, at concentrations present in the colon after a human therapeutic dosage of senna, rhein inhibited cell proliferation via a mechanism that seems to involve directly the MAP kinase pathway. Finally, rhein prevents the DNA damage probably via an anti‐oxidant mechanism.     

Identification of aminosuccinyl residues in peptides by second-derivative ultraviolet spectrometry

A second derivative ultraviolet spectrophotometric method for direct identification of aminosuccinyl peptides is described. The advantages and limits of this procedure have been assessed on the basis of results of five model peptides including the aminosuccinyl moiety.     

Interaction of Partially Purified Phytotoxins from Phytophthora cactorum on Apple Cell Plasma Membrane

The ‘crude’ filtrate (CF) of Phytophthora cactorum containing phytotoxin(s), having some properties similar to the toxins isolated from other Phytophthora species, was processed by three steps (acetone precipitation, dialysis and gel filtration chromatography). The CF fractions corresponding to the progressive steps of purification were tested for phytotoxicity on tomato seedlings and for activity on cell trans-membrane electrical potential (Em) of susceptible and resistant apple rootstocks (Malus domestica). The fractions (F4), obtained from chromatography on Sephadex G50 fine and eluted in the zone corresponding to a molecular weight of 15 ± 2 kD, induced a specific alteration on susceptible apple cell membranes. These metabolites, even though incompletely purified, are able to induce a high and specific activity on susceptible apple cell Em only, not on resistant ones. As a consequence, they may be of potential use in screening for insensitive cells.     

Interconverting flavanone glucosides and other phenolic compounds in Lippia salviaefolia Cham. ethanol extracts

Four interconverting flavanone glycosides [(2R)- and (2S)-3′,4′,5,6-tetrahydroxyflavanone 7-O-β-d-glucopyranoside, and (2R)- and (2S)-3′,4′,5,8-tetrahydroxyflavanone 7-O-β-d-glucopyranoside], in addition to eight known flavonoids [naringenin, asebogenin, sakuranetin, 6-hydroxyluteolin 7-O-β-d-glucoside, (2R)- and (2S)-eriodictyol 7-O-β-d-glucopyranoside, aromadendrin and phloretin], three phenylpropanoid glycosides [forsythoside B, alyssonoside and verbascoside] and the epoxylignan lariciresinol 4′-O-β-d-glucopyranoside were isolated and identified in the EtOH extract of the aerial parts of Lippia salviaefolia Cham. The phytochemical study herein was guided by preliminary antioxidant tests, namely, β-carotene protection and 2,2-diphenyl-1-picrylhydrazyl (DPPH) free radical scavenging activity. The crude extracts, their active fractions and the isolated compounds were assayed against intracellular reactive oxygen species (ROS) and human embryonic kidney HEK-293 and human melanoma M14 cancer cell growth. Aromadendrin and phloretin were able to counteract elevation of ROS induced by the oxidant t-butylhydroperoxide (t-BOOH) in HEK-293 cells, whereas phloretin strongly protected HEK-293 cells from ROS damage at 1 μM. Additionally, phloretin exhibited a significant growth inhibitory effect at 20–40 μM in both HEK-293 and M14 cells and induced a concentration dependent apoptosis at 20 μM in M14 cells, suggesting a selective action towards malignant cells. Due to their equilibria, the four interconverting flavanone glycosides were studied using 1D and 2D NMR, HPLC–CD–PDA and HRMS analyses.