Cardamom Extract Alleviates the Oxidative Stress, Inflammation and Apoptosis Induced during Acetaminophen-Induced Hepatic Toxicity via Modulating Nrf2/HO-1/NQO-1 Pathway.

Acetaminophen (APAP) is the most extensively used and safest analgesic and antipyretic drug worldwide; however, its toxicity is associated with life-threatening acute liver failure. Cardamom (CARD), a sweet, aromatic, commonly used spice, has several pharmacological actions. In the current study, we tried to explore the chemical composition and the hepato-protective effect of ethanolic aqueous extract of CARD to mitigate APAP-induced hepatic toxicity and elucidate its underlying mechanism of action. MATERIAL AND METHODS: Aqueous CARD extract was subjected to LC-TOF-MS analysis to separate and elucidate some of its components. In vivo animal experiments involved five groups of animals. In the normal and cardamom groups, mice were administered either saline or CARD (200 mg/kg), respectively, orally daily for 16 days. In the APAP group, the animals were administered saline orally daily for 15 days, and on the 16th day, animals were administered APAP (300 mg/kg) IP for the induction of acute hepatic failure. In the CARD 200 + APAP group, mice were administered CARD (200 mg/kg) for 15 days, followed by APAP on the 16th day. RESULTS: The aqueous extract of CARD showed several compounds, belonging to polyphenol, flavonoids, cinnamic acid derivatives and essential oil components. In the in vivo investigations, APAP-induced impaired liver function, several histopathological alterations, oxidative stress and inflammatory and apoptotic status signified severe hepatic failure. Whereas, pretreatment with the CARD extract prior to APAP administration diminished serum levels of the hepatic function test and augmented Nrf2 nucleoprotein and HO-1 and NQO-1. CARD down-regulated MDA, inflammatory mediators (IL-1ß, IL-6, TNF-α and NF-κB) and apoptotic markers (caspase 3 and 9 and Bax) and amplified the activities of SOD, catalase, GSH-Px and GSH-R in hepatic tissue samples. CONCLUSION: CARD extract mitigated the hepatic toxicity induced by APAP. The underlying mechanism of action of such hepato-protective action may be through upregulation of the Nrf2/HO-1/NQO-1 pathway with subsequent alleviation of the oxidative stress, inflammation and apoptosis induced by APAP. Many of the compounds identified in the CARD extract could be attributed to this pharmacological action of the extract.

Clinical Characteristics and Outcome of Candidemia: Experience from a Tertiary Referral Center in Saudi Arabia.

Background: Candida bloodstream infections cause significant excess morbidity and mortality in the health-care setting. There is limited evidence regarding Candida species causing invasive infections in Saudi Arabia. Objective: To identify Candida species causing bloodstream infection and determine the clinical outcome and factors associated with mortality in a tertiary center in Saudi Arabia. Materials and Methods: This retrospective study included all cases of positive blood culture for Candida in patients admitted to King Abdulaziz Medical City, a tertiary care center in Riyadh, Saudi Arabia, between January 1, 2013 and June 30, 2019. Results: A total of 532 patients with candidemia were identified (male: 55.4%; mean age: 54 ± 26.2 years). The most common Candida species isolated was Candida albicans (26.7%), followed by Candida glabrata (22.7%), Candida parapsilosis (22.2%), and Candida tropicalis (18.4%). Non-albicans candidemia was more common in patients with diabetes (76.7%; P = 0.0560), neutropenia (89.8%; P = 0.0062), recent exposure to fluconazole (85.7%; P = 0.0394), and active chemotherapy (83.1%; P = 0.0128). In non-albicans, susceptibility to fluconazole varied from 95.9% with C. tropicalis to 41.5% with C. parapsilosis; nonetheless, all species were highly susceptible to echinocandins. The overall 30- and 90-day mortality rates were 39.9% and 56.4%, respectively. The mortality rate was nonsignificantly higher with non-albicans species at 30 days (41.2% vs. 35.9%; P = 0.2634) and 90 days (58.2% vs. 51.4%; P = 0.1620). Conclusion: This study found a changing pattern in the Candida species causing bloodstream infections and an epidemiological shift toward more non-albicans Candida species in Saudi Arabia.

Physical activity and sedentary behaviors among active college students in Kuwait relative to gender status.

OBJECTIVES: To determine the prevalence of physical activity (PA), sedentary behaviors (SB) and sleep duration and the associations between those variables among Kuwaiti Physical Education (PE) College students. METHODS: A total of 418 participants (220 females) were randomly selected from the Basic Education College in the Public Authority for Applied Education and Training in Kuwait. Body weight and height were measured, and body mass index (BMI) was calculated. A validated questionnaire was used to assess lifestyle behaviors. RESULTS: Based on BMI classification, the prevalence of overweight or obesity among males (34.9%) was significantly higher than that of females (16.7%). However, the mean (SD) of body fat percentage using bioelectrical impedance analysis was 21.3 (9.0) for males and 32.3 (7.7) for females. The proportions of highly active (> 1,200 METs-min/week) males (85.9%) and females (64.3%) were significantly (p < 0.005) different. The participants exceeding 3 hours of screen viewing time/day and insufficient sleep duration (< 7 hours/night) were 76.8% and 65.1%, respectively, with no gender differences. Logistic regression, adjusted for confounders, showed that highly active PE students had a higher proportion of SB. However, neither sleep duration nor body fat percentage exhibited any significant difference relative to PA. CONCLUSIONS: Despite having high PA, Kuwaiti PE College students had a high prevalence of SB and insufficient sleep. Gender differences were found in PA but not in screen time or sleep duration. Efforts toward reducing SB and insufficient sleep among Kuwaiti PE College students are needed to reduce unhealthy lifestyle behaviors and promote health and well-being.

Structure-Activity Relationships of Potent, Targeted Covalent Inhibitors That Abolish Both the Transamidation and GTP Binding Activities of Human Tissue Transglutaminase.

Human tissue transglutaminase (hTG2) is a multifunctional enzyme. It is primarily known for its calcium-dependent transamidation activity that leads to formation of an isopeptide bond between glutamine and lysine residues found on the surface of proteins, but it is also a GTP binding protein. Overexpression and unregulated hTG2 activity have been associated with numerous human diseases, including cancer stem cell survival and metastatic phenotype. Herein, we present a series of targeted covalent inhibitors (TCIs) based on our previously reported Cbz-Lys scaffold. From this structure-activity relationship (SAR) study, novel irreversible inhibitors were identified that block the transamidation activity of hTG2 and allosterically abolish its GTP binding ability with a high degree of selectivity and efficiency (kinact/KI > 105 M-1 min-1). One optimized inhibitor (VA4) was also shown to inhibit epidermal cancer stem cell invasion with an EC50 of 3.9 µM, representing a significant improvement over our previously reported "hit" NC9.

Transglutaminase 2 modulation of NF-κB signaling in astrocytes is independent of its ability to mediate astrocytic viability in ischemic injury.

Transglutaminase 2 (TG2) is a multifunctional protein that can contribute to cell death and cell survival processes in a variety of disease contexts. Within the brain, TG2 has been shown to promote cell death in ischemic injury when expressed in astrocytes (Colak and Johnson, 2012). However, the specific functions and characteristics of astrocytic TG2 that mediate this effect are largely unknown. Therefore, the goal of this study was to investigate the role of astrocytic TG2 in mediating cellular viability processes in the context of ischemic injury, with a specific focus on its contributions to intracellular signaling cascades. We show that, in response to oxygen/glucose deprivation (OGD), acute lentiviral-mediated knockdown of TG2, as well as inhibition with an irreversible TG2 inhibitor, enhances cell survival. We also show that TG2 depletion increases nuclear factor-κB (NF-κB) signaling, whereas inhibition reduces NF-κB activity. Despite its clear contribution to NF-κB signaling, however, TG2 modulation of NF-κB signaling is not likely to be a major contributor to its ability to mediate astrocytic viability in this context. Overall, the results of this study provide insight into the role of TG2 in astrocytes and suggest possible avenues for future study of the relationship between astrocytic TG2 and ischemic injury.

Inhibition or ablation of transglutaminase 2 impairs astrocyte migration.

Astrocytes play numerous complex roles that support and facilitate the function of neurons. Further, when there is an injury to the central nervous system (CNS) they can both facilitate or ameliorate functional recovery depending on the location and severity of the injury. When a CNS injury is relatively severe a glial scar is formed, which is primarily composed of astrocytes. The glial scar can be both beneficial, by limiting inflammation, and detrimental, by preventing neuronal projections, to functional recovery. Thus, understanding the processes and proteins that regulate astrocyte migration in response to injury is still of fundamental importance. One protein that is likely involved in astrocyte migration is transglutaminase 2 (TG2); a multifunctional protein expressed ubiquitously throughout the brain. Its functions include transamidation and GTPase activity, among others, and previous studies have implicated TG2 as a regulator of migration. Therefore, we examined the role of TG2 in primary astrocyte migration subsequent to injury. Using wild type or TG2-/- astrocytes, we manipulated the different functions and conformation of TG2 with novel irreversible inhibitors or mutant versions of the protein. Results showed that both inhibition and ablation of TG2 in primary astrocytes significantly inhibit migration. Additionally, we show that the deficiency in migration caused by deletion of TG2 can only be rescued with the native protein and not with mutants. Finally, the addition of TGFß rescued the migration deficiency independent of TG2. Taken together, our study shows that transamidation and GTP/GDP-binding are necessary for inhibiting astrocyte migration and it is TGFß independent.

The complex role of transglutaminase 2 in glioblastoma proliferation.

BACKGROUND: Glioblastomas (GBMs) are a heterogeneous group of primary brain tumors. These tumors are resistant to therapeutic interventions and invariably recur after surgical resection. The multifunctional protein transglutaminase 2 (TG2) has been shown to promote cell survival in a number of different tumors. There is also evidence that TG2 may be a pro-survival factor in GBMs. However, the roles that TG2 plays in facilitating GBM survival and proliferation have not yet been clearly delineated . METHODS: The functions of TG2 are often cell- and context-specific. Therefore, in this study we examined the ability of TG2 to facilitate GBM proliferation using colony formation assays and 5-ethynyl-2'-deoxyuridine (EdU) incorporation in several different GBM cell lines as well as neurospheres derived from patient tumors representing the 3 major subtypes of GBM tumors (mesenchymal, proneural, and classical) and maintained in the absence of serum. TG2 knockdown or selective TG2 inhibitors were used to modulate TG2 expression and activity. RESULTS: We show that TG2 plays differential roles in the proliferative process depending on the cell type. In most, but not all, GBM models TG2 plays a crucial role in the proliferative process, and some but not all TG2 inhibitors were highly effective at reducing proliferation in a large subset of the GBM models. CONCLUSION: Our results show that TG2 plays an important-but notoriously context-specific-role in GBM cell biology. Nonetheless, as future studies unravel the genetic "fingerprints" that make TG2 inhibitors effective, this information could be exploited to develop TG2 inhibitors into personalized GBM therapies.

Simultaneous analysis of enzyme structure and activity by kinetic capillary electrophoresis-MS.

To enable the detection of protein conformational isomers, their enzymatic activity and their inhibition in a single experiment, we developed a method based on kinetic capillary electrophoresis coupled on-line with UV detection and ion mobility mass spectrometry (CE-UV-IM-MS). Kinetic CE-UV separated protein conformers and monitored their interconversion dynamics in solution. Ion mobility mass spectrometry analyzed the conformer sizes, exact molecular weights, and structures of an enzyme and of its substrates, inhibitors and corresponding products. This coupled CE-UV-IM-MS system allowed the simultaneous, real-time observation of the effect of small-molecule inhibitors on both the conformational distribution and enzymatic activity of the human tissue transglutaminase TG2. By expanding mass spectrometry profiling of enzymatic reactions beyond proteins and substrates to include protein dynamics, CE-UV-IM-MS opens a new avenue for the modulation and regulation of cellular functions, drug development and protein engineering.

Inhibitors of tissue transglutaminase.

Tissue transglutaminase (TG2) catalyzes the cross-linking of proteins by the formation of isopeptide bonds between glutamine (Gln) and lysine (Lys) side chains. Although TG2 is essential for the stabilization of the extracellular matrix, its unregulated activity has been implicated in celiac disease, fibrosis, and cancer metastasis, among other disorders. Given the importance and range of TG2-related pathologies, recent work has focused on the development of potent and selective inhibitors against TG2. In this review, we present the latest and most noteworthy irreversible and reversible inhibitors of TG2, and offer perspectives for the design of future inhibitors, in the hope that lead compounds with therapeutic potential may soon be discovered.

Acyl transfer mechanisms of tissue transglutaminase.

Tissue transglutaminase (TG2) is a calcium-dependent enzyme that catalyses several acyl transfer reactions. The most biologically relevant of these involve protein-bound Gln residues as an acyl-donor substrate, and either water or a primary amine as an acyl-acceptor substrate. The former leads to deamidation of Gln to Glu, whereas the latter leads to transamidation, typically resulting in protein cross-linking when the amine substrate is a protein-bound Lys residue. In this review, we present an overview of over fifty years of mechanistic studies that have led to our current understanding of TG2-mediated hydrolysis and transamidation.