Role of anthraquinones in Cassia occidentalis induced hepato-myo-encephalopathy.

ETHNOPHARMACOLOGICAL RELEVANCE: The different plant parts of Cassia occidentalis Linn, (CO) such as root, leaves, seeds and pods have traditionally been used in multifarious medicines for the treatment of dysentery, diarrhea, constipation, fever, eczema, cancer and venereal diseases. MATERIALS AND METHODS: A systematic search of literature has been done in books and scientific databases like Science Direct, Pubmed, Google Scholar and Scopus etc. These sources were used to compile, analyze and review the information regarding the phytochemistry, toxicology and mechanism of toxicity of CO. The various references on this subject are cited in our review ranging from 1956 to 2019. RESULTS: Unintentional exposure of CO causes serious pathological condition in children, known as hepato-myo-encephalopathy (HME). The toxicity after CO consumption is associated with the presence of anthraquinones (AQs), a class of secondary plant metabolites. These AQs at high concentrations are known to cause detrimental effects on essential vital organs such as liver, kidney, spleen, brain, muscle and reproductive organs. The animal studies in rodent models as well as clinical investigations have clearly revealed that CO toxicity is associated with enhanced hepatotoxicity serum markers (ALT, AST, and LDH) and presence of necrotic lesions in liver. Furthermore, CO also causes vacuolization in muscle tissue and increases the level of CPK which is a prominent muscle damage marker. Apart from these target organs, CO consumption also causes neuronal damage via disturbing the levels of different proteins such as (GFAP and b-tubulin III). The mechanistic studies show that AQs present in CO have the potential to disturb the cellular homeostasis via binding to DNA, increasing the production ROS and showing inhibitory effects on essential enzymes etc. Therefore, AQs have been observed to be the primary culprit agents contributing to the toxicity of CO in children and animals. CONCLUSION: Despite its therapeutic potential, CO consumption can be detrimental if consumed in high amounts. A thorough analysis of literature reveals that AQs are the primary factors contributing to toxicity of CO seeds. Exposure to CO seeds causes HME, which is a serious life threatening condition for the malnourished children from lower strata. Multiple mechanisms are involved in the CO induced HME in patients. Lack of appropriate diagnostic measures and a poor understanding of the CO toxicity mechanism in humans and animals complicate the clinical management of CO poisoning subjects. Therefore, development of point of care diagnostic kits shall help in early diagnosis & suitable management of CO poisoning.

Emodin inhibited NADPH-quinone reductase competitively and induced cytotoxicity in rat primary hepatocytes.

Consumption of Cassia occidentalis (CO) seeds, a ubiquitously distributed weed plant, is responsible for a pathological condition known as hepato-myo-encephalopathy (HME). The toxicity of CO seeds is largely attributed to the presence of anthraquinones (AQs). Here, we report that Emodin, a CO anthraquinone, inhibits the enzymatic activity of NADPH-Quinone reductase, which is an intracellular enzyme fundamentally involved in the detoxification of quinone containing compounds. Emodin binds to the active site of the enzyme and acts as a competitive inhibitor with respect to 2, 6-Dichlorophenolindophenol, a known substrate of NADPH-Quinone reductase. Moreover, our in-vitro study further revealed that Emodin was cytotoxic to primary rat hepatocytes.

N-acetyl-cysteine in combination with celecoxib inhibits Deoxynivalenol induced skin tumor initiation via induction of autophagic pathways in swiss mice.

Deoxynivalenol is a trichothecene mycotoxin which naturally contaminates small grain, cereals intended for human and animal consumption. Investigations for dermal toxicity of DON has been needed and highlighted by WHO. Previous studies on dermal toxicity suggest that DON has DNA damaging potential leading to skin tumor initiation in mice skin. However, considering its toxicological manifestations arising after dermal exposure, strategies for its prevention/protection are barely available in literatute. Collectively, our study demonstrated that N-acetylcysteine (NAC), precursor of glutathione, significantly alters the genotoxic potential of DON. Further NAC in combination with Celecoxib (CXB) inhibits tumor growth by altering antioxidant status and increasing autophagy in DON initiated Swiss mice. Despite the broad spectrum use of CXB, its use is limited by the concerns about its adverse effects on the cardiovascular system. Serum parameters and histology analysis revealed that CXB (2 mg) when applied topically for 24 weeks did not impart any cardiovascular toxicity which could be because skin permeation potential of CXB was quite low when analyzed through HPLC analysis. Although the anticancer effects of CXB and NAC have been studied, however, the combination of NAC and CXB has yet not been explored for any cancer treatment. Therefore our observations provide additional insights into the therapeutic effects of combinatorial treatment of CXB and NAC against skin tumor prevention. This approach might form a novel alternative strategy for skin cancer treatment as well as skin associated toxicities caused by mycotoxins such as DON. This combinatorial approach can overcome the limitations associated with the use of CXB for long term as topical application of the same seems to be safe in comparison to the oral mode of administration.

Emodin inhibited NADPH-quinone reductase via competitive mode of inhibition and induced cytotoxicity in rat primary hepatocytes.

Consumption of Cassia occidentalis (CO) seeds, a ubiquitously distributed weed plant, is responsible for a pathological condition known as hepato-myo-encephalopathy (HME). The toxicity of CO seeds is largely attributed to the presence of anthraquinones. Here, we report that Emodin, a CO anthraquinone, inhibits the enzymatic activity of NADPH-Quinone reductase, which is an intracellular enzyme fundamentally involved in the detoxification of quinone containing compounds. Emodin binds to the active site of the enzyme and acts as a competitive inhibitor with respect to 2, 6-Dichlorophenolindophenol, a known substrate of NADPH-Quinone reductase. Moreover, our in-vitro study further revealed that Emodin was cytotoxic to primary rat hepatocytes.

Methylenecyclopropyl glycine, not pesticide exposure as the primary etiological factor underlying hypoglycemic encephalopathy in Muzaffarpur, India.

Some districts of Bihar, especially Muzzaffarpur district, have been known to be affected by annual outbreak, called locally as Acute Encephalitis Syndrome (AES) which became one of the major health concerns in Bihar, due to its high fatality and complications. Several hypotheses like bat virus, heat stroke, pesticide exposure and the presence of a compound - methylenecyclopropyl glycine (MCPG) in Litchi have been proposed by different investigators for AES. When the investigators examined the symptoms, signs and the epidemiological data, bat virus and heat stroke hypothesis were ruled out. Two major hypotheses which remain in question were the exposure to pesticides or MCPG present in litchi. Therefore, this study was designed to check the presence of both in the Muzzaffarpur samples of ripe and semi ripe litchi fruits. The fruit cover of ripe and semi ripe litchi showed the traces of Malathion (0.18-0.19 µg/g) and p'-p'-DDT (0.022-0.023 µg/g), while no pesticide residues were detected in the pulp of ripe or semi ripe litchi thereby ruling out the possibility of pesticide exposure in children of Muzzaffarpur. However, MCPG was detected in the pulp of semi ripe (0.57 µg/g) and ripe litchi fruits (0.19 µg/g). Further, when the human condition was simulated in animals, there was deprivation in body weight and glucose levels in starved litchi seed dosed rats, causing hypoglycemia. These results suggest that the cause of hypoglycemic encephalopathy in Muzzaffarpur is related to the consumption of semi ripe and ripe litchi fruits by undernourished children.

Interaction of anthraquinones of Cassia occidentalis seeds with DNA and Glutathione.

Consumption of Cassia occidentalis (CO) seeds has been associated with the hepatomyoencephalopathy (HME) in children. Recently, we have characterized the toxic anthraquinones (AQs) such as Emodin, Rhein, Aloe-emodin, Chrysophanol and Physcion in CO seeds and detected these moieties in the bio fluids of CO poisoning cases. As AQs were detected in the serum of HME patients, their interaction with key biomolecules including protein, DNA and glutathione (GSH) is imperative. In this regard, we have previously reported the interaction of these AQs with serum albumin protein and their subsequent biological effects. However, the interaction of these AQs with DNA and GSH remained unexplored. In the present work, we have studied the binding of these AQs of CO seeds with DNA and GSH by fluorescence spectroscopy, UV-vis spectral analysis, molecular docking, and biochemical studies. Results indicated a higher binding affinity for Emodin (Ka = 3.854 × 104 L mol-1 S-1), Aloe-emodin (Ka = 0.961 × 104 L mol-1 S-1) and Rhein (Ka = 0.034 × 104 L mol-1 S-1) towards calf thymus DNA may be associated with their higher cytotoxicity. Alternatively, Physcion and Chrysophanol which showed less cytotoxicity in our earlier studies exhibited very low DNA binding. The binding pattern of all these AQs is consistent with the in-silico data. Absorption spectroscopy studies indicated the possible formation of GSH conjugate with Aloe-emodin and Physcion. Further biochemical measurement of GSH and GSSG (Glutathione disulfide) following incubation with AQs indicated that Aloe-emodin (28%) and Rhein (30%) oxidizes GSH to GSSG more as compared to other AQs. Taken together, these results suggest that the higher cytotoxicity of Rhein, Emodin and Aloe-emodin may be attributed to their potent DNA and GSH binding affinity.

A novel function of TLR4 in mediating the immunomodulatory effect of Benzanthrone, an environmental pollutant.

Our prior studies have reported that Benzanthrone (BA) manifests inflammatory responses in the spleen of Balb/c mice. The present investigation was carried out to study the impact of BA on macrophages, which are the primary scavenger cells in the body that act as a connecting link between innate and adaptive immunity. Parenteral administration of BA (daily for one week) to mice resulted in enhanced levels of nitric oxide (NO) and overexpression of inflammatory markers (COX-2, MMP-9 and PGE-2) in macrophages; however the level of MHC class-I and MHC class-II receptors were down regulated. Further, the potential membrane receptor targets (TLRs) of BA and its interaction with TLRs was investigated using computational methods. Professional phagocytes play pivotal roles in sensing bacteria through pathogen-associated molecular patterns (PAMPs) by various pathogen recognition receptors (PRRs), including Toll-like receptors (TLRs). Several studies have implicated these TLRs in the amplification of the inflammatory responses, however the fundamental role played by TLRs in mediating the inflammation associated with xenobiotics is still obscure and not understood. From the in silico analysis, it was evident that BA showed the highest binding affinity with TLR4 as compared to other TLRs. The western blotting studies confirmed that BA exposure indeed upregulated the expression of TLR 4, 5 and 9. Moreover, the downstream signaling cascade proteins of TLRs such as myeloid differentiation primary response protein-88 (MyD88), IL-1 receptor associated kinase (IRAK-1), and TNFR-associated factor (TRAF-6) were found to be enhanced in the BA treated groups. It was also observed that BA treatment increased the expression of ICAM-1, p-Lyn, p-Syk, p-PI3-K, IP3, PLC-γ, cAMP and Ca+2 influx, which are known to play a critical role in TLR mediated inflammation. Earlier we found that toxic effects of BA in spleen were mediated by oxidative stress which was partially neutralized by NAC exposure. Hereby, we report that NAC treatment in conjunction with BA attenuated the expression of BA induced TLR4, as well as the inflammatory markers such as COX2 and p-NFkB in macrophages. These findings demonstrated the critical role of TLRs in the regulation of the BA-induced inflammation.

Molecular Modeling and Docking Study to Elucidate Novel Chikungunya Virus nsP2 Protease Inhibitors.

Chikungunya is one of the tropical viral infections that severely affect the Asian and African countries. Absence of any suitable drugs or vaccines against Chikungunya virus till date makes it essential to identify and develop novel leads for the same. Recently, nsP2 cysteine protease has been classified as a crucial drug target to combat infections caused by Alphaviruses including Chikungunya virus due to its involvement viral replication. Here in, we investigated the structural aspects of the nsP2 protease through homology modeling based on nsP2 protease from Venezuelan equine encephalitis virus. Further, the ligands were virtually screened based on various pharmacological, ADME/Tox filters and subjected to docking with the modeled Chikungunya nsP2 protease using AutoDock4.2. The interaction profiling of ligand with the protein was carried out using LigPlot(+). The results demonstrated that the ligand with PubChem Id (CID_5808891) possessed highest binding affinity towards Chikungunya nsP2 protease with a good interaction profile with the active site residues. We hereby propose that these compounds could inhibit the nsP2 protease by binding to its active site. Moreover, they may provide structural scaffold for the design of novel leads with better efficacy and specificity for the nsP2 protease.

Evaluation extracellular matrix-chitosan composite films for wound healing application.

The present study describes the preparation of extracellular matrix (ECM; from porcine omentum) based chitosan composite films for wound dressing applications. The films were prepared by varying the ECM content, whereas, the amount of chitosan was kept constant. The interactions amongst the components of the films were analyzed by FTIR and XRD studies. The films were thoroughly characterized for surface hydrophilicity, moisture retention capability, water vapor permeability, mechanical and biocompatibility. FTIR study indicated that both chitosan and ECM were present in their native form and did not lose their activity. XRD analysis suggested composition dependent change in the crystallinity of the films. The mechanical properties suggested that the composite films had sufficient properties to be used for wound dressing applications. An increase in the ECM content resulted in better hydrophilicity of the films and hence better the moisture retention capacity and retardant water vapor transmission rate property of the composite films. The films were found to be biocompatible to both blood and adipose tissue derived stem cells. In gist, the prepared films may be explored as wound dressing materials.