3-Acetyl coumarin alleviate neuroinflammatory responses and oxidative stress in aluminum chloride-induced Alzheimer's disease rat model.

Alzheimer's disease (AD) is a neurodegenerative disorder that impairs mental ability and interrupts cognitive function. Heavy metal exposure like aluminum chloride is associated with neurotoxicity linked to neuro-inflammation, oxidative stress, accumulation of amyloid plaques, phosphorylation of tau proteins associated with AD like symptoms. The objective of the present investigation was to assess the effect 3-acetyl coumarin (3AC) in a rat model of AD. Preliminary screening was performed with SWISS ADME to check for the bioavailability of 3-AC and likeness score which proved favorable. 3-AC docked against Caspase 3, NF-κß and tau protein kinase I exhibited good binding energies. Male rats were divided into six groups (n = 5). AlCl3 (100 mg/kg BW) was administered for 28 days before starting treatment to induce AD. Normal control rats received vehicle. Treatment groups received 10, 20 and 30 mg/kg 3-AC for 28 days. Rivastigmine (2 mg/kg) was the standard. Behavioral tests (EPM, MWM) were performed at 7-day intervals throughout study period. Rats showed improved spatial memory and learning in treatment groups during behavioral tests. Rats were euthanized on day 28. Inflammatory markers (IL-1ß, IL-16 and TNFα) exhibited significant improvement (p < 0.001) in treated rats. Oxidative stress enzymes (SOD, CAT, GSH, MDA) were restored. Caspase3 and NF-κß quantified through qRT-PCR also decreased significantly (p < 0.001) when compared to disease control group. Levels of acetyl cholinesterase, dopamine and noradrenaline were also restored in treated rats significantly (p < 0.001). 3-AC treatment restored neuroprotection probably because of anti-inflammatory, anti-oxidant and anti-cholinesterase potential; hence, this can be considered a promising therapeutic potential alternative.

Quercetin-loaded chitosan nanoparticles ameliorate adjuvant-induced arthritis in rats by regulating anti-oxidant enzymes and downregulating pro- and inflammatory cytokines.

Rheumatoid arthritis (RA) is an inflammatory condition and associated with the symmetrical synovitis of the joints and cause joint pain. The use of anti-rheumatic drugs is associated with many adverse effects. Quercetin, an important polyphenolic flavonoid, possess anti-inflammatory and anti-rheumatic effects. Quercetin use is limited due to poor absorption and bioavailability. Nanomedicines are used for the targeted drug delivery, hence it reduces the adverse effects of the drug. Based upon these factors, quercetin-loaded chitosan nanoparticles (Q-NPs) were prepared by solvent evaporation method, characterized and their better anti-rheumatic effect with mechanistic insights was validated in Freund's complete adjuvant (FCA)-induced arthritic rats along with safety studies. The animals were divided into five groups, each containing 5 animals. Group I was normal control, group II was arthritic control, while groups III, IV and V were administered with quercetin (15 mg/Kg) and Q-NPs (10 and 20 mg/Kg), respectively. The reduction in ankle diameter, serum oxidative stress markers as well as pro- and inflammatory cytokines, e.g., tumor necrosis factor (TNFα), interleukin (IL-6) were determined. The prepared Q-NPs showed hydrodynamic size of 83.9 nm, polydispersity index of 0.687, entrapment efficiency 90.5% as well as no interaction between quercetin and chitosan in Fourier transform infrared spectroscopy (FTIR). A significant reduction (p < 0.001) in ankle diameter was observed after administration of high-dose Q-NPs (4.32 ± 0.14 cm to 5.13 ± 0.62 cm). There was also reduction (p < 0.001) in levels of TNFα and IL-6 following high-dose Q-NPs (72.56 ± 2.30 and 308.19 ± 11.5 pg). The effect on biochemical tests, hematological parameters and oxidative stress parameters was also found to be significant. Histopathological changes of kidney, liver and ankle also confirmed the anti-rheumatic effect of high-dose Q-NPs. The study concludes that administration of Q-NPs (20 mg/Kg) may be used for the treatment of FCA-induced RA in rats.

Pterostilbene reduces the progression of atopic dermatitis via modulating inflammatory and oxidative stress biomarkers in mice.

Atopic dermatitis (AD) is one of the most prevalent chronic skin inflammatory disorders requiring continuous treatment and care. Pterostilbene (PTN) belongs to stilbene and is a polyphenolic compound of natural origin. It is similar to resveratrol and has analogous anti-inflammatory, anti-oxidant, and anti-carcinogenic characteristics. This study was intended to evaluate the effect of PTN against atopic dermatitis. The disease was induced by sensitization with 2,4-dinitrochlorobenzene (DNCB) in mice. The standard control group (SCG) received topical 0.1% tacrolimus (TC), whereas three other treatment groups received daily topical PTN at 0.2, 0.6, and 1% w/w for 28 days. Dermatitis scoring, ear thickness, and body weight of animals were weekly determined while other parameters were assessed at the termination of the experiment. PTN reduced the ear weight, skin thickness, and the weight and size of thymus glands and spleen in comparison with diseased animals. PTN also reduced the elevated immunoglobulin E (IgE) level and blood inflammatory cells in diseased mice. The histopathological findings showed a decreased epidermal thickness in PTN-treated groups. Moreover, treatment with PTN improved the amount of oxidative stress markers in the skin of the diseased mice. The expressions of IL-4, IL-6, TNF-α, and NF-κB in the skin of diseased mice were also reduced by PTN. This study concludes that PTN ameliorated the symptoms of atopic dermatitis through the reduction of inflammation, oxidative damage, and inflammatory cytokines in the skin of diseased animals. Therefore, PTN must be further investigated for the treatment of AD complications and other inflammatory skin disorders.

Deletion of the phosphatase INPP5E in the murine retina impairs photoreceptor axoneme formation and prevents disc morphogenesis.

INPP5E, also known as pharbin, is a ubiquitously expressed phosphatidylinositol polyphosphate 5-phosphatase that is typically located in the primary cilia and modulates the phosphoinositide composition of membranes. Mutations to or loss of INPP5E is associated with ciliary dysfunction. INPP5E missense mutations of the phosphatase catalytic domain cause Joubert syndrome in humans-a syndromic ciliopathy affecting multiple tissues including the brain, liver, kidney, and retina. In contrast to other primary cilia, photoreceptor INPP5E is prominently expressed in the inner segment and connecting cilium and absent in the outer segment, which is a modified primary cilium dedicated to phototransduction. To investigate how loss of INPP5e causes retina degeneration, we generated mice with a retina-specific KO (Inpp5eF/F;Six3Cre, abbreviated as retInpp5e-/-). These mice exhibit a rapidly progressing rod-cone degeneration resembling Leber congenital amaurosis that is nearly completed by postnatal day 21 (P21) in the central retina. Mutant cone outer segments contain vesicles instead of discs as early as P8. Although P10 mutant outer segments contain structural and phototransduction proteins, axonemal structure and disc membranes fail to form. Connecting cilia of retInpp5e-/- rods display accumulation of intraflagellar transport particles A and B at their distal ends, suggesting disrupted intraflagellar transport. Although INPP5E ablation may not prevent delivery of outer segment-specific proteins by means of the photoreceptor secretory pathway, its absence prevents the assembly of axonemal and disc components. Herein, we suggest a model for INPP5E-Leber congenital amaurosis, proposing how deletion of INPP5E may interrupt axoneme extension and disc membrane elaboration.

Therapeutic potential of Ceratonia siliqua extract for the management of asthma and hypertension.

The current work was performed to explore the pharmacological mechanisms involved in the management of asthma and hypertension along with the safety profile of the Ceratonia siliqua (C. siliqua/Carob) pods. The bronchorelaxant, vasorelaxant, and cardioselective activities of C. siliqua pods were investigated using isolated rabbit tracheal, aortic, and paired atrial fragments on the Power lab data acquisition system. Normotensive rats were used to study antihypertensive activity. The plant extract and its fractions relaxed the carbachol-induced contraction in the tracheal fragments and shifted the concentration-response curve of carbachol towards the right confirming the muscarinic receptor antagonist activity. The relaxation of phenylephrine-induced contraction in an aortic fragment by the extract showed α- adrenergic blocking activity. Furthermore, the extract produced a cardio-selective response in the paired atria and decreased the blood pressure in anesthetized normotensive rats. The plant extract proved to be non-toxic in oral acute and chronic toxicity studies and did not demonstrate any sign of histopathological lesions. These results suggested that the plant extract was non-toxic and could be used in the management of lifetime therapies of respiratory and cardiovascular disorders without any unwanted effects.

Comparative Study of Nano-liposome and Nano-niosome for Delivery of Achillea Millefolium Essential Oils: Development, Optimization, Characterization and Their Cytotoxicity Effects on Cancer Cell Lines and Antibacterial Activity.

Nanoencapsulation of essential oils (EOs) in drug delivery systems leads to their capability of improving their solubility, stability, and bioavailability of them. The aim of this study was preparation, optimization, and characterization of nano-liposomes/nano-niosomes containing Achillea millefolium essential oils (A. millefolium EOs) and comparison of their properties. In the experimental study, characteristics of nanoparticles including size, zeta potential, Fourier Transform Infrared Spectroscopy (FTIR), % encapsulation efficiency (EE%) and the release amount of essential oils from nano-liposome or niosome were assessed using different techniques. Then to determine cell viability at different concentrations, the MTT assay was used. Also, the dilution method was used to determine the minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of antimicrobial agents. The optimized formulations provided potential advantages, including an appropriate nano-size scale, and a negative charge, and also showed a continuous drug release behavior, which successfully encapsulated essential oil with high entrapment efficiency. In terms of size and release amount, nano-niosome had superiority to nano-liposome with smaller size and also slower release but nano-liposome could encapsulate essential oils in a higher percentage compared to nano-niosome. Also, there was a significant difference between the toxicity of encapsulated EOs and free EOs in terms of viability (P<0.05). In addition, the antimicrobial effect of liposomal and niosomal EO was greater than free EO. In conclusion, the designed nano-based systems were determined as promising lipid-based nano-carriers for essential oil delivery that proffered a novel, high potential therapy for breast cancer and favorable antimicrobial effects.

Methotrexate-loaded biodegradable nanoparticles exert anti-arthritic effect by downregulating pro-inflammatory cytokines in Freund's complete adjuvant-induced arthritic rats.

Methotrexate (MTX), the first-line drug for the treatment of rheumatoid arthritis (RA), can cause considerable toxicity, which limits effective dosage regimens. Moreover, it has rapid clearance, which leads to poor patient compliance. To mitigate such challenges, this study aimed to validate the use of MTX-loaded chitosan nanoparticles (NPs) in treating Freund's complete adjuvant (FCA) arthritis in rats. Healthy Wistar rats (n = 30) were divided into five groups. The first group served as healthy control, while the second group served as arthritic control. Group 3 was administered methotrexate, while groups 4 and 5 were MTX-loaded NP-treated groups. NPs were prepared by solvent evaporation method and characterized by zeta size, potential, polydispersity index (PDI), and Fourier-transform infrared spectroscopy. NPs were 190 nm in size, and PDI was 0.25, confirming the uniform distribution of NPs. A significant increase in paw thickness was noted up to the 21st day of the study, which was reversed by a high dose of MTX-loaded NPs. MTX NPs significantly reduced the level of pro-inflammatory markers, including TNF-α and IL-6, along with improving control of oxidative stress biomarkers. The findings of biochemical, haematological, radiological, and histopathological investigations further confirmed amelioration of necrosis and cellular infiltration. It can be concluded that MTX-loaded chitosan NPs are promising candidates for treating FCA-induced arthritis in a rat model.

Pathogenic role of cytokines in COVID-19, its association with contributing co-morbidities and possible therapeutic regimens.

The Covid-19, a threatening pandemic, was originated from China in December 2019 and spread quickly to all over the world. The pathogenesis of coronavirus is linked with the disproportionate response of the immune system. This involves the systemic inflammatory reaction which is characterized by marked pro-inflammatory cytokine release commonly known as cytokine release storm (CRS). The pro inflammatory cytokines are involved in cascade of pulmonary inflammation, hyper coagulation and thrombosis which may be lethal for the individual. That's why, it is very important to have understanding of pro inflammatory cytokines and their pathological role in SARS-CoV-2. The pathogenesis of Covid is not the same in every individual, it can vary due to the presence of pre-existing comorbidities like suffering from already an inflammatory disease such as rheumatoid arthritis (RA), inflammatory bowel disease (IBD), chronic obstructive pulmonary disease (COPD), an immune-compromised patients suffering from Diabetes Mellitus (DM) and Tuberculosis (TB) are more vulnerable morbidity and complications following COVID-19. This review is particularly related to COVID-19 patients having comorbidity of other inflammatory diseases. We have discussed the brief pathogenesis of COVID-19 and cytokines release storm with reference to other co-morbidities including RA, IBD, COPD, DM and TB. The available therapeutic regimens for COVID-19 including cytokine inhibitors, anti-viral, anti-biotic, bronchodilators, JAK- inhibitors, immunomodulators and anti-fibrotic agents have also been discussed briefly. Moreover, newly emerging medicines in the clinical trials have also been discussed which are found to be effective in treating Covid-19.

Anticancer, Cardio-Protective and Anti-Inflammatory Potential of Natural-Sources-Derived Phenolic Acids.

Phenolic acids (PAs) are one of the utmost prevalent classes of plant-derived bioactive chemicals. They have a specific taste and odor, and are found in numerous medicinal and food plants, such as Cynomorium coccineum L., Prunus domestica (L.), and Vitis vinifera L. Their biosynthesis, physical and chemical characteristics and structure-activity relationship are well understood. These phytochemicals and their derivatives exert several bioactivities including but not limited to anticancer, cardioprotective, anti-inflammatory, immune-regulatory and anti-obesity properties. They are strong antioxidants because of hydroxyl groups which play pivotal role in their anticancer, anti-inflammatory and cardioprotective potential. They may play significant role in improving human health owing to anticarcinogenic, anti-arthritis, antihypertensive, anti-stroke, and anti-atherosclerosis activities, as several PAs have demonstrated biological activities against these disease during in vitro and in vivo studies. These PAs exhibited anticancer action by promoting apoptosis, targeting angiogenesis, and reducing abnormal cell growth, while anti-inflammatory activity was attributed to reducing proinflammatory cytokines. Pas exhibited anti-atherosclerotic activity via inhibition of platelets. Moreover, they also reduced cardiovascular complications such as myocardial infarction and stroke by activating Paraoxonase 1. The present review focuses on the plant sources, structure activity relationship, anticancer, anti-inflammatory and cardioprotective actions of PAs that is attributed to modulation of oxidative stress and signal transduction pathways, along with highlighting their mechanism of actions in disease conditions. Further, preclinical and clinical studies must be carried out to evaluate the mechanism of action and drug targets of PAs to understand their therapeutic actions and disease therapy in humans, respectively.

Pterostilbene improves CFA-induced arthritis and peripheral neuropathy through modulation of oxidative stress, inflammatory cytokines and neurotransmitters in Wistar rats.

Pterostilbene is a stilbene flavonoid that occurs naturally in various plants as well as produced by genetic engineering. It exhibits anti-inflammatory, analgesic, anti-oxidant and neuroprotective activities. This research was aimed to determine the potential of pterostilbene against arthritis and peripheral neuropathy in Complete Freund's Adjuvant (CFA) induced arthritis. Rat hind paw was injected with 0.1 ml CFA to induce arthritis. Standard control animals received oral methotrexate (3 mg/kg/week). Pterostilbene at 12.5, 25 and 50 mg/kg was given orally to different groups of arthritic rats from day 7-28 for 21 days. Pterostilbene significantly reduced paw diameter and retarded the decrease in body weight of arthritic rats. It profoundly (p < 0.05-0.0001) reduced lipid peroxidation and nitrites, while increased superoxide dismutase (SOD) in the liver tissue. Pterostilbene treatment significantly (p < 0.0001) reduced TNF-α and IL-6 levels. Pterostilbene markedly improved (p < 0.05-0.001) motor activity and showed analgesic effect in arthritic rats at 25 and 50 mg/kg as compared to disease control rats. Furthermore, it notably (p < 0.05-0.0001) increased SOD activity, nitrites, noradrenaline and serotonin levels in the sciatic nerve of arthritic rats. Treatment with pterostilbene also ameliorated the CFA-induced pannus formation, cartilage damage and synovial hyperplasia in the arthritic rat paws. It is determined from the current study that pterostilbene was effective in reducing CFA-induced arthritis in rats through amelioration of oxidative stress and inflammatory mediators. It was also effective to treat peripheral neuropathy through modulation of oxidative stress and neurotransmitters in sciatic nerves.

Effect of packaging material and oxygen concentration on quality of Ghavoot during the storage.

Ghavoot is an Iranian traditional food product that prepared by a combination of several types of plant seeds mixed with sugar. The lack of appropriate packaging caused Ghavoot exposed to environment conditions, which leads to oxidation of this product and reduce its nutritional value and marketability. In this study, different types of packaging materials including nylon, the Polyester/ Aluminum/ polyethylene (PET/AL/PE) and the Polyester /Aluminum / Low-Density Polyethylene (PET/AL/LDPE) with different concentrations of oxygen inside the packaging (zero, 5 and 21%) were used to maintain quality properties of Ghavoot. The results showed that samples stored in the PET/Al/LDPE packaging under vacuum, had fewer moisture changes during storage compared with other treatments, as a result, the least changes in the color parameters of Ghavoot occurred. Increasing the concentration of oxygen inside the packaging resulted in higher peroxide, anisidine and totox values as well as the higher total acidity of the Ghavoot's oil. Keeping Ghavoot in the three-layer PET/Al/LDPE pouches under vacuum condition, caused the lipid oxidation to be delayed during the storage. Results of sensory properties showed that increasing the oxygen concentration inside the packaging caused the average score for product flavor to decrease as a result of rancidity development.

Iranian superwomen's career experiences: a qualitative study.

BACKGROUND: Superwoman refers to the identity of a woman who performs several important roles simultaneously and full-time, such as being a wife, mother, and homemaker while holding a job. This study aims to examine the career experiences of Iranian superwomen who maintained their mental health while holding multiple roles. METHODS: Data for this qualitative study were collected via semi-structured interview and analyzed using conventional content analysis. The study participants were 12 multi-role women from different occupations in Tabriz, North West of Iran selected through purposive and theoretical sampling. The women's mental health status was assessed using Mental Health Continuum-Short Form questionnaire before participating in the research. RESULTS: The results were classified into three main categories. The first category included Underlying Factors of Job Experience with six subcategories, including Limited job opportunities for women, Educational context (mindset shaped in the parental home), Commitment or freedom in obtaining a job and its continuation, Personality traits, and Non-cognitive skills (emotional intelligence and spiritual intelligence); the second category included the adopted strategies to solve career problems with two subcategories: effective strategies, including the benefits of having a mindset of abundance and having a vision and strategic planning, and ineffective strategies, including the benefits of having a mindset of scarcity; and the third category included Perceived Consequences of employment with two subcategories: psychological consequences and social consequences, with both subcategories including some positive and negative further subcategories. Moreover, a conceptual relationship between the main categories and the subcategories was evident. CONCLUSIONS: The content obtained not only supports the findings about the experiences of multi-role women in cultures outside Iran, but also points to the unique aspects of Iranian superwomen's life experiences and narratives. The findings of this study can help us perceive the real career experiences from the perspective of Iranian professional women with multiple roles.

Genotoxic and cytotoxic assessment of sitagliptin and simvastatin alone and in combination.

Type 2 Diabetes Mellitus (T2DM) patients are at high risk of Coronary Heart Disease (CHD) and need a global therapeutic intervention. A fixed-dose combination prescription medication containing anti-diabetic drug (Sitagliptin) and lipid lowering (Simvastatin) has recently been approved. Present study was designed to explore the potential synergistic toxic effects of sitagliptin and simvastatin at cellular level. MTT assay revealed the potential synergistic cytotoxic effect whereas Comet assay spotlighted the genotoxicity. MTT assay conducted on Vero cell lines revealed no significant change in proliferative activity upon treatment with simvastatin but cell survival percentage (CSP) decreased upon treatment with sitagliptin (51% at 1000µg/mL). However, combination of both drugs exhibited a better survival percentage except highest dose combination (1000:500µg/mL) which augmented antiproliferative effects rendering CSP 71.6%. The genotoxic assay spotted that Simvastatin produced less damage to DNA with the threshold of 500µg/ml whereas Sitagliptin significantly damage above the 250µg/mL, However, combination of drugs produced lesser damage than Sitagliptin alone. The findings concluded a non-genotoxic combination of sitagliptin and simvastatin which possess a least cytotoxic potential suggesting the safe use of the combination both in T2DM and CHD.

Pharmacological, Phytochemical and histopathological basis of Conyza bonariensis in the potential management of diabetes mellitus.

To evaluate the anti-diabetic potential of aqueous methanolic extract of Conyza bonariensis amongst the Wistar rats. Phytochemical and High Performance Liquid Chromatography (HPLC) analyses of phenols and flavonoids were examined. The plant extract (250 and 500mg/kg/day) was explored for its anti-hyperglycemic effect for 14 days in normoglycemic and alloxan-induced diabetic rats using the oral glucose tolerance test (OGTT). HPLC analyses demonstrated the composition of the plant extract as gallic acid, cinnamic acid, quercetin, p-coumaric acid and syringic acid. The blood glucose concentrations in experimental diabetic as well as non-diabetic rats significantly decreased with doses 250 and 500 mg/kg in OGTT. Moreover, the significant drop in fasting glucose level was observed following 14 days of therapy. It also ameliorated the serum cholesterol, total protein, low and high density lipoproteins, glycosylated hemoglobin A1C and serum amylase with respect to untreated rats suffering from diabetes. There appeared to be no significant alteration with regard to body weight amongst the treated rats. The plant extract revamped the pancreatic islets of Langerhans and abridged alloxan-induced degenerative changes in the liver. It can be concluded that Conyza bonariensis extract has a pronounced hypoglycemic effect on diabetes due to the presence of phytochemicals.

Berberis lycium Royle. extracts attenuate inflammation and modulates hyperglycemia in alloxan induced diabetic rats.

Berberis lycium Royle (Berberidaceae) is traditionally used for the treatment of diabetes mellitus. Present study was conducted to determine the antioxidant, antidiabetic and anti-inflammatory effects of aqueous and methanolic whole plant extracts. Total phenolic contents were determined by Folin-ciocalteu method whereas antioxidant activity was determined by 2,2-diphenyl-1-picryl hydrazyl (DPPH) method. In vitro anti-diabetic activity was determined using alpha amylase assay. Acute hypoglycemic activity was investigated on normoglycemic rats. Sub-acute anti-diabetic effects were investigated in alloxan induced diabetic rats for 14 days. Methanolic extract exhibited 183.5±1 mg/g Gallic acid equivalent (GAE) phenolic contents. The methanolic extract exhibited an IC50 of 242µg/mL and 37.26 mg/mL in antioxidant and alpha amylase inhibitory assays respectively. Administration of methanolic extract in normoglycemic rats exhibited significant anti-hyperglycemic effect at 90 and 120 min. Methanolic extract (500 mg/kg extract) significantly reduced blood glucose at day 14. Methanolic extract (500 mg/kg) significantly reduced the concentration of tumor necrosis factor (TNF-α) and interleukin (IL-6) along with reduction in total cholesterol and triglyceride levels in diabetic rats. Administration of methanol extract also improved the hepatic markers. The study suggested that the methanolic extract possessed antidiabetic effect that might be attributed to its alpha amylase, antioxidant and anti-inflammatory potential.

Phytochemical, anti-inflammatory, anti-nociceptive and cytotoxic basis for the use of Haloxylon stocksii.

A halophytic plant, Haloxylon stocksii, is used to treat various inflammatory disorders traditionally. The present study was carried out to investigate the phytochemical parameters, anti-inflammatory, analgesic and cytotoxic potential of the whole plant extracts of H. stocksii. The plant powder was standardized for pharmacognostic parameters. It was extracted with methanol followed by chloroform, ethyl acetate and water to prepare respective fractions. Total phenolic and flavonoid contents in the extract and fractions were estimated. The anti-inflammatory potential was determined through carrageenan-induced rat paw edema model. Centrally acting analgesic activity was assessed through the hot plate method. MTT assay was used to assess the viability of Human umbilical and human hepatocyte carcinoma cell lines upon exposure to plant extract/fractions. Chloroform fraction showed the highest phenolic while ethyl acetate exhibited a maximum flavonoids content. The plant ethyl acetate fraction exhibited highest percentage inhibition of paw edema and maximum analgesic activity at 500 mg/kg dose. The plant methanolic extract and fractions showed dose dependent cytotoxic activity. The present study concludes that the extracts of H. stocksii may be effective and safe against acute inflammatory response and pain at therapeutic concentrations.

C8ORF37 Is Required for Photoreceptor Outer Segment Disc Morphogenesis by Maintaining Outer Segment Membrane Protein Homeostasis.

C8ORF37 is a causative gene for three different clinical forms of incurable retinal degeneration. However, the completely unknown function of C8ORF37 limits our understanding of the pathogenicity of C8ORF37 mutations. Here, we performed a comprehensive phenotypic characterization of a C8orf37 KO mouse line, generated using CRISPR/Cas9 technology. Both C8orf37 KO male and female mice exhibited progressive and simultaneous degeneration of rod and cone photoreceptors but no non-ocular phenotypes. The major ultrastructural feature of C8orf37 KO photoreceptors was massive disorganization of the outer segment (OS) membrane discs starting from the onset of disc morphogenesis during development. At the molecular level, the amounts of multiple OS-specific membrane proteins, including proteins involved in membrane disc organization, were reduced, although these proteins were targeted normally to the OS. Considering the distribution of C8ORF37 throughout the photoreceptor cell body, the normal structure of the KO photoreceptor connecting cilium, and the absence of defects in other ciliary organs of the KO mice, our findings do not support the previous notion that C8ORF37 was a ciliary protein. Because C8ORF37 is absent in the photoreceptor OS, C8ORF37 may participate in the secretory pathway of OS membrane proteins in the photoreceptor cell body and thus maintain the homeostasis of these proteins. This study established a valid animal model for future therapeutic studies of C8ORF37-associated retinal degeneration. This study also shed new light on the role of C8ORF37 in photoreceptors and on the pathogenic mechanism underlying retinal degeneration caused by C8ORF37 mutations.SIGNIFICANCE STATEMENT Inherited retinal degeneration is a group of incurable conditions with poorly understood underlying molecular mechanisms. We investigated C8ORF37, a causative gene for three retinal degenerative conditions: retinitis pigmentosa, cone-rod dystrophy, and Bardet-Biedl syndrome. C8ORF37 encodes a protein with no known functional domains and thus its biological function is unpredictable. We knocked out the C8ORF37 ortholog in mice, which resulted in a retinal phenotype similar to that observed in patients. We further demonstrated that C8ORF37 is required for photoreceptor outer segment disc formation and alignment, a process that is critical for photoreceptor function and survival. This study advances our understanding of the pathogenesis of retinal degeneration and establishes a valuable mouse model for future therapeutic development.

Antioxidant and alpha amylase inhibitory activities of Fumaria officinalis and its antidiabetic potential against alloxan induced diabetes.

Fumaria officinalis belongs to family papaveraceae and is traditionally used to treat hypertension, hepatitis and diabetes. The current study was conducted to evaluate in vitro and in vivo antidiabetic activity of Fumaria officinalis. Aerial parts of the plant were sequentially extracted with n-hexane, chloroform, methanol and water. Phytochemical analysis was carried out on all extracts. Antioxidant activity was determined by 2,2-diphenyl-1-picryl hydrazyl (DPPH) inhibition method. In vitro alpha-amylase inhibitory activity was performed on all extracts by using dinitrosalicylic acid. Effect of aqueous and methanolic extracts of F. officinalis on blood glucose was evaluated in normo-glycaemic rats and alloxan induced diabetic rats. Glimepiride 0.2 mg/kg was used as standard therapy in diabetic rats. Results showed that methanolic extract exhibited the maximum percentage inhibition of DPPH (86.30%) and alpha-amylase inhibition (94.01%) at 500 µg/ml and 16 mg/ml concentration respectively. Administration in normo-glycaemic rats did not show any significant decrease in blood glucose level at 500 and 750 mg/kg dosage. Aqueous and methanolic extracts exhibited a significant hypoglycaemic effect (p˂0.05) at all doses. A significant increase in the body weight and an improvement in liver and kidney function tests of diabetic rats were observed. These extracts also reduced the damage to the cells of glomeruli, interstitial inflammation, necrosis of tubular cells and thrombosis in the kidney, the enlargement of sinusoids and steatosis in the liver of diabetic rats. This study concludes that F. officinalis may have antidiabetic potential possibly due to its antioxidant and alpha-amylase inhibitory activities.

In vitro and in vivo anti-arthritic evaluation of Polystichum braunii to validate its folkloric claim.

Medicinal plants are playing an imperative role in the therapy for treating various chronic ailments including arthritis. The present study was focused on finding in-vitro and in-vivo anti-arthritic potential of P. braunii roots. In vitro protein denaturation, membrane stabilization and anti-trypsinase assays were carried out to demonstrate anti-arthritic activity of the extracts. Furthermore, the extracts exerting promising in vitro anti-arthritic potential were tested orally at 150, 300 and 600mg/kg/day against formaldehyde induced arthritis in Wistar rats. The methanolic, aqueous and ethyl acetate extracts of the plant revealed noteworthy in vitro anti-arthritic activities while mitigating formaldehyde induced paw edema in dose dependent manner. Methanolic and aqueous extracts showed the highest inhibition (p<0.05) of paw edema, arthritic indices, reduced elevated level of platelets and leukocytes while increasing hemoglobin and body weight of arthritic rats. Anti-arthritic activity of the plant extracts may be due to inhibition of protein denaturation and lysosomal membrane stabilization. The plant exhibited good anti-arthritic potential.

The guanine nucleotide exchange factor Arf-like protein 13b is essential for assembly of the mouse photoreceptor transition zone and outer segment.

Arf-like protein 13b (ARL13b) is a small GTPase that functions as a guanosine nucleotide exchange factor (GEF) for ARL3-GDP. ARL13b is located exclusively in photoreceptor outer segments (OS) presumably anchored to discs by palmitoylation, whereas ARL3 is an inner segment cytoplasmic protein. Hypomorphic mutations affecting the ARL13b G-domain inactivate GEF activity and lead to Joubert syndrome (JS) in humans. However, the molecular mechanisms in ARL13b mutation-induced Joubert syndrome, particularly the function of primary cilia, are still incompletely understood. Because Arl13b germline knockouts in mouse are lethal, we generated retina-specific deletions of ARL13b in which ARL3-GTP formation is impaired. In mouse retArl13b-/- central retina at postnatal day 6 (P6) and older, outer segments were absent, thereby preventing trafficking of outer segment proteins to their destination. Ultrastructure of postnatal day 10 (P10) central retArl13b-/- photoreceptors revealed docking of basal bodies to cell membranes, but mature transition zones and disc structures were absent. Deletion of ARL13b in adult mice via tamoxifen-induced Cre/loxP recombination indicated that axonemes gradually shorten and outer segments progressively degenerate. IFT88, essential for anterograde intraflagellar transport (IFT), was significantly reduced at tamArl13b-/- basal bodies, suggesting impairment of intraflagellar transport. AAV2/8 vector-mediated ARL13b expression in the retArl13b-/- retina rescued ciliogenesis.