Patient-derived colon epithelial organoids reveal lipid-related metabolic dysfunction in pediatric ulcerative colitis.

Ulcerative colitis (UC) is associated with epithelial metabolic derangements which exacerbate gut inflammation. Patient-derived organoids recapitulate complexities of the parent tissue in health and disease; however, whether colon organoids (colonoids) model the metabolic impairments in the pediatric UC epithelium is unclear. Here, we developed colonoid lines from pediatric patients with endoscopically active UC, inactive UC, and those without endoscopic or histologic evidence of colon inflammation (non-IBD controls) to interrogate functional metabolic differences in the colon epithelia. We demonstrate that colonoids from active UC patients exhibit hypermetabolic features and cellular stress, specifically during differentiation. Hypermetabolism in differentiating active UC colonoids was driven, in part, by increased proton leak, and supported by enhanced glycolytic capacity and dysregulated neutral lipid accumulation. Transcriptomic and pathway analyses indicated a role for PPAR-α in lipid-induced hypermetabolism in aUC colonoids, which was validated by PPAR-α activation in non-IBD colonoids. Accordingly, limiting neutral lipid accumulation in active UC colonoids through pharmacological inhibition of PPAR-α induced a metabolic shift towards glucose utilization, suppressed hypermetabolism and chemokine secretion, and improved markers of cellular stress and epithelial differentiation. Taken together, we reveal a role for lipid-related metabolic dysfunction in the pediatric UC epithelium and support the advancement of colonoids as a preclinical human model for testing epithelial-directed therapies against such metabolic dysfunction.

Neuroprotective mechanism of trans,trans-Farnesol in an ICV-STZ-induced rat model of Alzheimer's pathology.

BACKGROUND: Alzheimer's disease (AD) is a prominent cause of dementia, resulting in neurodegeneration and memory impairment. This condition imposes a considerable public health burden on both patients and their families due to the patients' functional impairments as well as the psychological and financial constraints. It has been well demonstrated that its aetiology involves proteinopathy, mitochondriopathies, and enhanced reactive oxygen species (ROS) generation, which are some of the key features of AD brains that further result in oxidative stress, excitotoxicity, autophagy, and mitochondrial dysfunction. OBJECTIVE: The current investigation was created with the aim of elucidating the neurological defence mechanism of trans,trans-Farnesol (TF) against intracerebroventricular-streptozotocin (ICV-STZ)-induced Alzheimer-like symptoms and related pathologies in rodents. MATERIALS AND METHODS: The current investigation involved male SD rats receiving TF (25-100 mg/kg, per oral) consecutively for 21 days in ICV-STZ-treated animals. An in silico study was carried out to explore the possible interaction between TF and NADH dehydrogenase and succinate dehydrogenase. Further, various behavioural (Morris water maze and novel object recognition test), biochemical (oxidants and anti-oxidant markers), activities of mitochondrial enzyme complexes and acetylcholinesterase (AChE), pro-inflammatory (tumor necrosis factor-alpha; TNF-α) levels, and histopathological studies were evaluated in specific brain regions. RESULTS: Rats administered ICV-STZ followed by treatment with TF (25, 50, and 100 mg/kg) for 21 days had significantly better mental performance (reduced escape latency to access platform, extended time spent in target quadrant, and improved differential index) in the Morris water maze test and new object recognition test models when compared to control (ICV-STZ)-treated groups. Further, TF treatment significantly restored redox proportion, anti-oxidant levels, regained mitochondrial capacities, attenuated altered AChE action, levels of TNF-α, and histopathological alterations in certain brain regions in comparison with control. In in silico analysis, TF caused greater interaction with NADH dehydrogenase and succinate dehydrogenase. CONCLUSION: The current work demonstrates the neuroprotective ability of TF in an experimental model with AD-like pathologies. The study further suggests that the neuroprotective impacts of TF may be related to its effects on TNF-α levels, oxidative stress pathways, and mitochondrial complex capabilities.

Thoracic and abdominal aortic alterations in dogs affected with systemic hypertension.

Aortic remodeling is the consequence of untreated systemic hypertension along with aortic dilatation as a marker for target organ damage in human literature. Therefore, the present study was planned to detect the changes in aorta at the level of aortic root via echocardiography, thoracic descending aorta via radiography and abdominal aorta via ultrasonography in healthy (n = 46), diseased normotensive (n = 20) and systemically hypertensive dogs (n = 60). The aortic root dimensions were measured at the level of aortic annulus, sinus of valsalva, sino-tubular junction and proximal ascending aorta via left ventricular outflow tract view of echocardiography. The thoracic descending aorta was subjectively assessed for any disparity in size and shape of aorta via lateral and dorso-ventral view of chest radiography. The abdominal aorta was assessed via left and right paralumbar window for calculating the aortic elasticity along with aortic and caudal venacaval dimensions to calculate the aortic-caval ratio. The aortic root measurements were dilated (p < 0.001) in systemically hypertensive dogs with a positive correlation (p < 0.001) with systolic blood pressure (BP). Thoracic descending aorta was also (p < 0.05) altered in the size and shape (undulation) of systemically hypertensive dogs. Abdominal aorta was markedly stiffened with reduced elasticity (p < 0.05) along with dilatation (p < 0.01) in hypertensive dogs. Also, there was a positive correlation (p < 0.001) of aortic diameters and aortic-caval ratio and negative correlation (p < 0.001) of aortic elasticity with systolic BP. Therefore, it was concluded that aorta could be considered as an important target organ damage of systemic hypertension in dogs.

Marine-derived Natural Products as Anticancer Agents.

Cancer is a deadly human disease on the rise due to changes in lifestyle, nutrition, and global warming. Cancer is characterized by uncontrolled, disordered, and undesired cell division. About 60% of cancer medicines approved by the FDA are made from natural ingredients. Intensive efforts over the last decade to better understand the vast chemical diversity provided by marine life have resulted in an intriguing "marine pipeline" of potential anticancer clinical and preclinical treatments. The molecular targets of marine products as anticancer drugs, as well as different reported compounds acting on distinct targets, are the topic of this review.

Methionine uptake via the SLC43A2 transporter is essential for regulatory T-cell survival.

Cell death, survival, or growth decisions in T-cell subsets depend on interplay between cytokine-dependent and metabolic processes. The metabolic requirements of T-regulatory cells (Tregs) for their survival and how these are satisfied remain unclear. Herein, we identified a necessary requirement of methionine uptake and usage for Tregs survival upon IL-2 deprivation. Activated Tregs have high methionine uptake and usage to S-adenosyl methionine, and this uptake is essential for Tregs survival in conditions of IL-2 deprivation. We identify a solute carrier protein SLC43A2 transporter, regulated in a Notch1-dependent manner that is necessary for this methionine uptake and Tregs viability. Collectively, we uncover a specifically regulated mechanism of methionine import in Tregs that is required for cells to adapt to cytokine withdrawal. We highlight the need for methionine availability and metabolism in contextually regulating cell death in this immunosuppressive population of T cells.

Sirtuin1 meditated modification of Notch1 intracellular domain regulates nucleolar localization and activation of distinct signaling cascades.

Notch signaling is involved in cell fate decisions in the development and maintenance of tissue homeostasis. Spatial regulation of the Notch1 intracellular domain (NIC1), has been shown to underpin signaling outcomes mediated by this receptor. We recently reported a putative Nucleolar Localization Sequence (NoLS) in NIC1. Here we investigate if the putative NoLS identified in NIC1 regulates localization in the nucleolus and anti-apoptotic activity. Confocal imaging of live cells expressing NIC1 or forms modified by deletion or site-directed mutagenesis established that the putative NoLS in NIC1 is required for nucleolar localization and regulated by the deacetylase Sirtuin1. Subsequent analysis of anti-apoptotic activity revealed signaling cascades linked to nucleolar localization. For this, etoposide and 4-Nitroquinoline 1-oxide, an inhibitor of topoisomerase-II and a UV mimetic drug respectively, were used as prototypic triggers of genomic damage in a mammalian cell line. While NIC1 blocked apoptosis regardless of its localization to the nucleoplasm or nucleolus, modifications of NIC1 which promoted localization to the nucleolus triggered a dependence on the nucleolar proteins fibrillarin and nucleolin for anti-apoptotic activity. Further, cells co-expressing NIC1 and Sirtuin1 (but not its catalytically inactive form), confirmed both spatial regulation and the switch to dependence on the nucleolar proteins. Finally, site-directed mutagenesis showed that the NoLS lysine residues are targets of Sirtuin1 activity. NIC1 mediated transcription is not similarly regulated. Thus, NIC1 localization to the nucleolus is regulated by Sirtuin1 modification of the lysine residues in NoLS and triggers a distinct signaling cascade involving nucleolar intermediates for anti-apoptotic activity.

S-nitrosoglutathione alleviates hyperglycemia-induced neurobehavioral deficits involving nitro-oxidative stress and aberrant monaminergic system.

The present study evaluated the protective role of S-nitrosoglutathione (GSNO) in preventing hyperglycemia-induced nitro-oxidative stress and alterations in monoaminergic system associated with neurobehavioral deficits in mice. Mice were subjected to diabetes by intraperitoneal injection of streptozotocin (40 mg/kg body weight) for 5 days, whereas GSNO (100 µg/kg body weight) was administered daily via oral route for 8 weeks. Diabetic mice showed deficits in neurobehavioral functions associated with memory, learning, anxiety and motor coordination. These neurobehavioral deficits observed in diabetic mice may be attributed to decrease in norepinephrine (NE), dopamine (DA), serotonin (5-HT) and increased monoamine oxidase (MAO) activity in cortex and hippocampus. Further, a significant increase in reactive oxygen species (ROS), protein carbonyls, nitrotyrosine (NT) and lipid peroxidation were observed in brain regions of diabetic animals suggesting increased nitro-oxidative stress. Hyperglycemia induced nitro-oxidative stress appears to involve reduction in redox ratio (GSH/GSSG) and enzymatic antioxidants; catalase (CAT) and superoxide dismutase (SOD) in cortex and hippocampus. However, GSNO supplementation was able to ameliorate alterations in monoaminergic system and nitro-oxidative stress in the brain regions thereby restoring neurobehavioural functions. These findings suggest GSNO as potential therapeutic molecule to prevent diabetic encephalopathy.

Notch1 Modulation of Cellular Calcium Regulates Mitochondrial Metabolism and Anti-Apoptotic Activity in T-Regulatory Cells.

As the major hub of metabolic activity and an organelle sequestering pro-apoptogenic intermediates, mitochondria lie at the crossroads of cellular decisions of death and survival. Intracellular calcium is a key regulator of these outcomes with rapid, uncontrolled uptake into mitochondria, activating pro-apoptotic cascades that trigger cell death. Here, we show that calcium uptake and mitochondrial metabolism in murine T-regulatory cells (Tregs) is tuned by Notch1 activity. Based on analysis of Tregs and the HEK cell line, we present evidence that modulation of cellular calcium dynamics underpins Notch1 regulation of mitochondrial homeostasis and consequently anti-apoptotic activity. Targeted siRNA-mediated ablations reveal dependency on molecules controlling calcium release from the endoplasmic reticulum (ER) and the chaperone, glucose-regulated protein 75 (Grp75), the associated protein Voltage Dependent Anion Channel (VDAC)1 and the Mitochondrial Calcium Uniporter (MCU), which together facilitate ER calcium transfer and uptake into the mitochondria. Endogenous Notch1 is detected in immune-complexes with Grp75 and VDAC1. Deficits in mitochondrial oxidative and survival in Notch1 deficient Tregs, were corrected by the expression of recombinant Notch1 intracellular domain, and in part by recombinant Grp75. Thus, the modulation of calcium dynamics and consequently mitochondrial metabolism underlies Treg survival in conditions of nutrient stress. This work positions a key role for Notch1 activity in these outcomes.

Ebstein's anomaly in a French bulldog.

A one and a half years old male French bulldog weighing 9.50 kg was presented with the history of inappetence, lethargy, abdominal distension and exercise intolerance since last 2 days. The physical examination was done which revealed normal physiological parameters including temperature, mucus membrane color and capillary refill time except palpable precordial thrills, jugular distension on palpation, tachycardia and systolic murmurs on auscultation. Electrocardiography (ECG) was done which depicted ectopic foci with atrioventricular junctional tachycardia and right ventricular enlargement involving very small inverted P waves, deep S waves in leads I, II, III and augmented vector foot (aVF) and splintered QRS complexes. The dog was undergone chest radiography that revealed right atrial enlargement, increased sternal contact of heart on lateral view and a bulge at 9:00 o'clock to 11:00 o'clock depicted right atrial enlargement on dorso-ventral view. Lastly, echocardiography was done to arrive at a diagnosis confirming the Ebstein's anomaly as a form of tricuspid valve dysplasia including apical displacement of tricuspid valve leaflets, division of right ventricle into atrialized and functional portions, increased displacement index, increased apex-mitral annulus to apex-tricuspid annulus ratio, severe right atrial dilatation and tricuspid regurgitation. The dog was medically treated with diuretics, angiotensin converting enzyme inhibitors and inotropes and the owner was advised to put the dog on low sodium diet for 2 weeks. The dog has resolved clinical signs of right sided heart affection; but, suddenly collapsed at home. The owner denied for the necropsy of dog.

Therapeutic potential of Allium Sativum against the Aß(1-40)-induced oxidative stress and mitochondrial dysfunction in the Wistar rats.

From the early stages of any neurodegenerative-disease mitochondrial functionality has been mortally extricated, though the exact timeline of these events is still unclear, it is likely to represent a progressive neurons-decline and cognitive-functions. Hence strategies suggested by herbal extract to restore mitochondrial functions may be a remedial approach to chronic neurodegenerative disorder like Alzheimer's disease (AD). This research was designed to evaluate if Aß1-40 induced oxidative stress and mitochondrial dysfunction could be inhibited by Allium Sativum (AS) supplementation. AD was induced by a single intra-hippocampal injection of Aß1-40 (5 µg/4 µl), while herbal supplementation was given orally (100, 250, 500 mg/kg body weight, daily) for 3 weeks. Morris water maze was used to assess cognitive function shows deficits in Aß1-40 treated animals, there is no significant alteration in locomotor function as examined by actophotometer. This was accompanied by enhancement in oxidative stress indicating by accentuated ROS and protein carbonyl levels. Concomitantly, decrease in activity of antioxidant enzymes was observed in diseased animals; as expressed by reduced superoxide-dismutase and catalase activity, as well as reduction in GSH levels and impaired mitochondrial functions. Medium dose of AS has been found effective in restoring the memory impairment along with antioxidant levels but high dose is more efficient as observed in the Aß1-40 treated rats. High dose of AS, on the other hand significantly ameliorates the mitochondrial-dysfunction in comparison to medium dose. Taken together, the findings reveal that AS reverses Aß1-40 induced brain alteration, it could be an efficient clinical mitigation action against AD growth.

Spatial regulation and generation of diversity in signaling pathways.

Signaling pathways orchestrate diverse cellular outcomes in the same tissue, spatially and temporally. These interactions, which are played out in micro-environments within cells and involve a relatively small number of core pathways, are the key to the development and function of multi-cellular organisms. How these outcomes are regulated has prompted interest in intracellular mechanisms that build diversity in signaling outcomes. This review specifically addresses spatial positioning of molecules as a means of enabling interactions and novel outcomes of signaling cascades. Using the Notch and Ras pathways as exemplars, we describe mechanisms that contribute to diverse signaling outcomes.

Nucleolar localization of the Notch4 intracellular domain underpins its regulation of the cellular response to genotoxic stressors.

Cell survival is one of the many cellular processes regulated by Notch family of proteins. A comparison of human breast cancer cell lines, which differ in the levels of endogenous Notch4, implicated the protein in regulating susceptibility to apoptosis triggered by genomic damage. In agreement with this observation, increased susceptibility to genotoxic damage was observed following siRNA ablations of Notch4 in two breast cancer cell lines. Further, overexpressing Notch4 intracellular domain (NIC4) tagged to GFP (NIC4-GFP), protected cells from apoptosis triggered by genotoxic drugs. In cells immune-stained for endogenous Notch4, protein was detected in the nucleolus and nucleoplasm, which was also confirmed by the co-localization of NIC4-GFP with RFP-tagged nucleolar proteins in breast cancer cells or the unrelated HEK cell line. Linking functional outcomes to nucleolar localization, NIC4-GFP protection from apoptosis, required the nucleolar proteins Nucleolin and Fibrillarin. Consistently, immunoprecipitation analysis revealed associations between nucleolar proteins-Nucleolin and Nucleophosmin-and Notch4. Microscopy-based biophysical analysis of live cells showed that nucleolar and nucleoplasmic pools of NIC4-GFP are mobile, with some sequestration of nucleolar NIC4-GFP pools. A nucleolar excluded form, NIC4_3RA-GFP, generated by site-directed mutagenesis of the nucleolar localization sequence in NIC4, could not protect from apoptosis triggered by genotoxic stressors. However, transcriptional activity or protection from apoptosis triggered by endoplasmic stress was comparable in cells expressing NIC4_3RA-GFP or NIC4-GFP. Together, the data show that nucleolar localization of NIC4 is critical for the regulation of genomic damage and may be uncoupled from its activities in the nucleoplasm. This study identifies intrinsic features of NIC4 that regulate signaling outcomes activated by the receptor by controlling its spatial localization.

Bacopa monnieri prevents colchicine-induced dementia by anti-inflammatory action.

Inflammation is considered as an early event in the development of Alzheimer's disease (AD) that precedes the formation of Aß plaques and neurofibrillary tangles. Therefore, strategies aimed at attenuating inflammation by phytochemicals may be a potential therapeutic intervention against AD. The present study was designed to evaluate if colchicine-induced inflammation and Aß production could be prevented by Bacopa monnieri (BM) supplementation. Dementia was induced by a single intracerebroventicular injection of colchicine (15 µg/5 µl), whereas, BM extract was administered orally (50 mg/kg body weight, daily) for 15 days. Assessment of cognitive functions using Morris water maze revealed deficits in colchicine administered animals. This was accompanied by significant increase in oxidative stress in terms of accentuated ROS and NO production. Expression of pro-inflammatory cytokines (IL-6, TNF-α) and chemokine (MCP-1) increased in the brain regions. Furthermore, COX-2 and iNOS expression also increased significantly in the brain regions of colchicine-administered animals. In addition, BACE-1 activity increased in the colchicine treated animals, which was accompanied by enhanced Aß production. On the other hand, BM supplementation was able to improve cognitive functions, suppress Aß formation by reducing BACE-1 activity. Inflammatory and oxidative stress markers were attenuated in the brain regions of BM supplemented animals. Taken together, the findings reveal that BM reverses colchicine-induced dementia by its anti-inflammatory and anti-oxidant action suggesting that it may be an effective therapeutic intervention to ameliorate progression of AD.

Intravaginal administration of metformin hydrochloride loaded cationic niosomes amalgamated with thermosensitive gel for the treatment of polycystic ovary syndrome: In vitro and in vivo studies.

BACKGROUND AND OBJECTIVE: Metformin hydrochloride (MTF-HCl) is extensively recommended by physicians for the treatment of polycystic ovary syndrome (PCOS). Mechanistically, MTF-HCl activates AMP-dependent kinase-α (AMPK-α) pathway to decrease the glucose production, enhances fatty acid oxidation and elevates the uptake of glucose in tissues. However, despite favourable physicochemical properties, oral administration of MTF-HCl is associated with impaired bioavailability (50-60%), lactic-acidosis and frequent dosing (500mg 2-3 times a day) in PCOS that ultimately influence the patient compliance. Therefore, in present investigation, MTF-HCl loaded unmodified and cationic small unilamellar niosomes were separately amalgamated with thermosensitive gel (MTF-HCl-SUNs-Gel and MTF-HCl-C-SUNs-Gel) for the treatment of PCOS through vaginal route of administration. METHODS AND RESULTS: MTF-HCl-SUNs and MTF-HCl-C-SUNs were separately prepared by reverse phase evaporation method. The nanovesicle size and zeta-potential of MTF-HCl-C-SUNs were measured to be 210.3±14.8-nm (P<0.05) and +8.7±2.7-mV (P<0.001), significantly higher than 198.5±20.3-nm and -16.6±3.9-mV of MTF-HCl-SUNs, respectively. Moreover, promising results of in vitro characterization parameters like gelation time, gelling temperature, viscosity analysis, percent mucoadhesiveness and drug release of MTF-HCl-C-SUNs-Gel and MTF-HCl-SUNs-Gel ensured the candidature of tailored gels for further in vivo investigations. In this way, treatment of PCOS rats under scheduled dose-dosage regimen with oral MTF-HCl solution, intravaginal MTF-HCl-SUNs-Gel and intravaginal MTF-HCl-C-SUNs-gel exhibited remarkable alterations, recruitment and development of normal follicles in addition to normalization of level of various hormones in PCOS. CONCLUSION: In conclusion, MTF-C-SUNs-Gel has paved the way for developing intravaginal dosage form of MTF-HCl for the treatment of PCOS.

Neuroprotective effects of Bacopa monnieri in experimental model of dementia.

Alzheimer disease (AD) is characterized by dementia that begins as mild short term memory deficit and culminates in total loss of cognitive and executive functions. The present study was conducted to evaluate the neuroprotective potential of Bacopa monnieri (BM), an Indian traditional medicinal plant effective against cognitive impairment, in colchicine-induced dementia. Intracerebroventricular administration of colchicine (15 µg/5 µl) induced cognitive impairment in rats as assessed by elevated plus maze. This was accompanied by a significant increase in oxidative stress in term of enhanced levels of lipid peroxidation and protein carbonyls. Concomitantly, decrease in activity of antioxidant enzymes was observed in colchicine treated animals. BM (50 mg/kg body weight) supplementation reversed memory impairment observed in the colchicine treated rats. BM administration attenuated oxidative damage, as evident by decreased LPO and protein carbonyl levels and restoration in activities of the antioxidant enzymes. The activity of membrane bound enzymes (Na(+)K(+) ATPase and AChE) was altered in colchicine treated brain regions and BM supplementation was able to restore the activity of enzymes to comparable values observed in controls. The results suggest therapeutic potential of BM in the treatment of AD associated cognitive decline.

Successful treatment of pulmonary paragonimiasis in a German shepherd dog with fenbendazole.

Pulmonary paragonimiasis is an important zoonotic disease reported from many parts of the world. It is an endemic problem in human population in north-eastern states of India. There seems no report of pulmonary paragonimiasis in canine population from India. The present case describes first report of pulmonary paragonimiasis in a female dog suggesting possibility of this fluke becoming established in canine population in the country. The dog revealed mild coughing with serous nasal discharge. Faecal sample revealed eggs of Paragonimus spp. Treatment with fenbendazole resulted in marked improvement as revealed by clinical signs and chest radiography.