Prohibitin 2 Is an Inner Mitochondrial Membrane Mitophagy Receptor.

The removal of unwanted or damaged mitochondria by autophagy, a process called mitophagy, is essential for key events in development, cellular homeostasis, tumor suppression, and prevention of neurodegeneration and aging. However, the precise mechanisms of mitophagy remain uncertain. Here, we identify the inner mitochondrial membrane protein, prohibitin 2 (PHB2), as a crucial mitophagy receptor involved in targeting mitochondria for autophagic degradation. PHB2 binds the autophagosomal membrane-associated protein LC3 through an LC3-interaction region (LIR) domain upon mitochondrial depolarization and proteasome-dependent outer membrane rupture. PHB2 is required for Parkin-induced mitophagy in mammalian cells and for the clearance of paternal mitochondria after embryonic fertilization in C. elegans. Our findings pinpoint a conserved mechanism of eukaryotic mitophagy and demonstrate a function of prohibitin 2 that may underlie its roles in physiology, aging, and disease.

MYC promotes tryptophan uptake and metabolism by the kynurenine pathway in colon cancer.

Tumors display increased uptake and processing of nutrients to fulfill the demands of rapidly proliferating cancer cells. Seminal studies have shown that the proto-oncogene MYC promotes metabolic reprogramming by altering glutamine uptake and metabolism in cancer cells. How MYC regulates the metabolism of other amino acids in cancer is not fully understood. Using high-performance liquid chromatography (HPLC)-tandem mass spectrometry (LC-MS/MS), we found that MYC increased intracellular levels of tryptophan and tryptophan metabolites in the kynurenine pathway. MYC induced the expression of the tryptophan transporters SLC7A5 and SLC1A5 and the enzyme arylformamidase (AFMID), involved in the conversion of tryptophan into kynurenine. SLC7A5, SLC1A5, and AFMID were elevated in colon cancer cells and tissues, and kynurenine was significantly greater in tumor samples than in the respective adjacent normal tissue from patients with colon cancer. Compared with normal human colonic epithelial cells, colon cancer cells were more sensitive to the depletion of tryptophan. Blocking enzymes in the kynurenine pathway caused preferential death of established colon cancer cells and transformed colonic organoids. We found that only kynurenine and no other tryptophan metabolite promotes the nuclear translocation of the transcription factor aryl hydrocarbon receptor (AHR). Blocking the interaction between AHR and kynurenine with CH223191 reduced the proliferation of colon cancer cells. Therefore, we propose that limiting cellular kynurenine or its downstream targets could present a new strategy to reduce the proliferation of MYC-dependent cancer cells.

Mitochondrial dynamics and inheritance during cell division, development and disease.

During cell division, it is critical to properly partition functional sets of organelles to each daughter cell. The partitioning of mitochondria shares some common features with that of other organelles, particularly in the use of interactions with cytoskeletal elements to facilitate delivery to the daughter cells. However, mitochondria have unique features - including their own genome and a maternal mode of germline transmission - that place additional demands on this process. Consequently, mechanisms have evolved to regulate mitochondrial segregation during cell division, oogenesis, fertilization and tissue development, as well as to ensure the integrity of these organelles and their DNA, including fusion-fission dynamics, organelle transport, mitophagy and genetic selection of functional genomes. Defects in these processes can lead to cell and tissue pathologies.

TLR9 and beclin 1 crosstalk regulates muscle AMPK activation in exercise.

The activation of adenosine monophosphate-activated protein kinase (AMPK) in skeletal muscle coordinates systemic metabolic responses to exercise1. Autophagy-a lysosomal degradation pathway that maintains cellular homeostasis2-is upregulated during exercise, and a core autophagy protein, beclin 1, is required for AMPK activation in skeletal muscle3. Here we describe a role for the innate immune-sensing molecule Toll-like receptor 9 (TLR9)4, and its interaction with beclin 1, in exercise-induced activation of AMPK in skeletal muscle. Mice that lack TLR9 are deficient in both exercise-induced activation of AMPK and plasma membrane localization of the GLUT4 glucose transporter in skeletal muscle, but are not deficient in autophagy. TLR9 binds beclin 1, and this interaction is increased by energy stress (glucose starvation and endurance exercise) and decreased by a BCL2 mutation3,5 that blocks the disruption of BCL2-beclin 1 binding. TLR9 regulates the assembly of the endolysosomal phosphatidylinositol 3-kinase complex (PI3KC3-C2)-which contains beclin 1 and UVRAG-in skeletal muscle during exercise, and knockout of beclin 1 or UVRAG inhibits the cellular AMPK activation induced by glucose starvation. Moreover, TLR9 functions in a muscle-autonomous fashion in ex vivo contraction-induced AMPK activation, glucose uptake and beclin 1-UVRAG complex assembly. These findings reveal a heretofore undescribed role for a Toll-like receptor in skeletal-muscle AMPK activation and glucose metabolism during exercise, as well as unexpected crosstalk between this innate immune sensor and autophagy proteins.

Multiresponsive Nanoscale Self-Assembly of Azurin-Elastin-like Polypeptide Fusion Protein for Enhanced Prostate Cancer Therapy.

A fusion protein composed of a bacterial protein, azurin, having antineoplastic properties and a thermally responsive structural cationic elastin-like protein (ELP), is designed, cloned, expressed, and purified. A simple method of inverse transition cycle (ITC) is employed to purify the fusion protein azurin-ELP diblock copolymer (d-bc). The molecular weight of the azurin-ELP fusion protein is ∼32 kDa. Further, its self-assembly properties are investigated. Interestingly, the engineered azurin-ELP d-bc in response to increasing temperature shows a dual-step phase separation into biofunctional nanostructures. Around the physiological temperature, azurin-ELP d-bc forms stable coacervates, which is dependent on the concentration and time of incubation. These coacervates are formed below the lower critical solubility temperature (LCST) of the ELP block at physiological temperature. Above LCST, i.e., 50-55°C, micelles of size ranging from 25 to 30 nm are formed. The cytotoxicity of azurin-ELP d-bc depends on the size of the coacervates formed and their cellular uptake at physiological temperature. Further, MTT assay of azurin-ELP d-bc in the cross-linked micelles prepared ex situ shows > six times higher killing of LNCaP cells than the unimeric form of azurin-ELP at 5 µM concentration. The flow cytometric results of these micelles at 20 µM concentration show ∼97% LNCaP cells in the apoptotic phase. Thus, azurin-ELP cross-linked micelles have enhanced potential for anticancer therapy due to their higher avidity.

A Retrospective Study to Evaluate Real-world Evidence of Azelnidipine in Indian Patients (REDEFINE Study).

INTRODUCTION: Azelnidipine, a selective calcium channel blocker, effectively lowers blood pressure (BP) and heart rate (HR) in hypertensive patients, as demonstrated in a retrospective real-world evidence (RWE) study in Indian patients. MATERIALS AND METHODS: This was a retrospective cohort study that included 882 patients aged 18 years or older who had been on azelnidipine treatment for the last 3 months for mild to moderate hypertension (HTN). A structured proforma was utilized to gather data from prescribing physicians to assess the efficacy of azelnidipine (8 and 16 mg) as monotherapy or in combination with other antihypertensive drugs. The primary endpoints of the study were to capture changes in systolic blood pressure (SBP) and diastolic BP (DBP) from baseline to the subsequent visits (4 and 12 weeks), while the secondary endpoints were to measure similar changes in the diabetic group and to estimate the proportion of patients achieving target BP of <130/80 mm Hg and <140/90 mm Hg, respectively. RESULTS: The overall mean reduction of systolic/diastolic BP from baseline to 12 weeks was 13.92/7.91 mm Hg (p-value < 0.0001). The mean reduction of systolic/diastolic BP from baseline to 12 weeks was 11.77/7.43 mm Hg (p-value < 0.0001) in newly diagnosed HTN patients, while in known cases of HTN, it was 16.50/8.48 mm Hg (p-value < 0.0001). In the diabetic group, the mean reduction was 15.35/8.69 mm Hg (p-value < 0.0001). Overall the study showed that in 44 (4.99%) and 408 (46.26%) patients, target BP of <130/80 mm Hg and <140/90 mm Hg, respectively was achieved. The mean change in HR from baseline was a reduction of 5.22 beats/minute. CONCLUSION: Azelnidipine can be an effective antihypertensive drug to treat mild to moderate HTN in Indian patients.

Engineered Recombinant EGFP-Azurin Theranostic Nanosystem for Targeted Therapy of Prostate Cancer.

Erythropoietin-producing hepatocellular (Eph) receptors and their ligands, ephrins, are the largest subfamily of receptor tyrosine kinases (RTKs) that have emerged as a new class of cancer biomarkers due to their aberrant expression in cancer progression. The activation of Eph receptors either due to their hyperexpression or via high affinity binding with their respective ephrin ligands initiates a cascade of signals that impacts cancer development and progression. In prostate cancer, the overexpression of the EphA6 receptor has been correlated with increased metastatic potential. Azurin, a small redox protein, is known to prevent tumor progression by binding to cell surface Eph receptors, inhibiting its autophosphorylation in the kinase domain and thereby disrupting Eph-ephrin signaling. Hence, a self-assembled, theranostic nanosystem of recombinant fusion protein his6EGFP-azu (80-128) was designed by conjugating enhanced green fluorescent protein (EGFP) with the C-terminal region of azurin. This design was inspired by the in silico binding study, where the analogue of ephrinA, his6EGFP-azu (80-128) showed higher binding affinity for the EphA6 receptor than the ephrinA ligands. The his6EGFP-azu (80-128) nanosystem which assembled as nanoparticles was tested for its ability to simultaneously detect and kill the prostate cancer cells, LNCaP. This was achieved by specifically targeting EphA6 receptors overexpressed on the cancer cell surface via C-terminal peptide, azu (80-128). Herein, we report antiproliferative, apoptotic, antimigratory, and anti-invasive effects of this nanosystem on LNCaP cells, while having no similar effects on EphA6 negative human normal lung cells, WI-38.

IndiGenomes: a comprehensive resource of genetic variants from over 1000 Indian genomes.

With the advent of next-generation sequencing, large-scale initiatives for mining whole genomes and exomes have been employed to better understand global or population-level genetic architecture. India encompasses more than 17% of the world population with extensive genetic diversity, but is under-represented in the global sequencing datasets. This gave us the impetus to perform and analyze the whole genome sequencing of 1029 healthy Indian individuals under the pilot phase of the 'IndiGen' program. We generated a compendium of 55,898,122 single allelic genetic variants from geographically distinct Indian genomes and calculated the allele frequency, allele count, allele number, along with the number of heterozygous or homozygous individuals. In the present study, these variants were systematically annotated using publicly available population databases and can be accessed through a browsable online database named as 'IndiGenomes' http://clingen.igib.res.in/indigen/. The IndiGenomes database will help clinicians and researchers in exploring the genetic component underlying medical conditions. Till date, this is the most comprehensive genetic variant resource for the Indian population and is made freely available for academic utility. The resource has also been accessed extensively by the worldwide community since it's launch.

Skp, Cullin, F-box (SCF)-Met30 and SCF-Cdc4-Mediated Proteolysis of CENP-A Prevents Mislocalization of CENP-A for Chromosomal Stability in Budding Yeast.

Restricting the localization of the histone H3 variant CENP-A (Cse4 in yeast, CID in flies) to centromeres is essential for faithful chromosome segregation. Mislocalization of CENP-A leads to chromosomal instability (CIN) in yeast, fly and human cells. Overexpression and mislocalization of CENP-A has been observed in many cancers and this correlates with increased invasiveness and poor prognosis. Yet genes that regulate CENP-A levels and localization under physiological conditions have not been defined. In this study we used a genome-wide genetic screen to identify essential genes required for Cse4 homeostasis to prevent its mislocalization for chromosomal stability. We show that two Skp, Cullin, F-box (SCF) ubiquitin ligases with the evolutionarily conserved F-box proteins Met30 and Cdc4 interact and cooperatively regulate proteolysis of endogenous Cse4 and prevent its mislocalization for faithful chromosome segregation under physiological conditions. The interaction of Met30 with Cdc4 is independent of the D domain, which is essential for their homodimerization and ubiquitination of other substrates. The requirement for both Cdc4 and Met30 for ubiquitination is specifc for Cse4; and a common substrate for Cdc4 and Met30 has not previously been described. Met30 is necessary for the interaction between Cdc4 and Cse4, and defects in this interaction lead to stabilization and mislocalization of Cse4, which in turn contributes to CIN. We provide the first direct link between Cse4 mislocalization to defects in kinetochore structure and show that SCF-mediated proteolysis of Cse4 is a major mechanism that prevents stable maintenance of Cse4 at non-centromeric regions, thus ensuring faithful chromosome segregation. In summary, we have identified essential pathways that regulate cellular levels of endogenous Cse4 and shown that proteolysis of Cse4 by SCF-Met30/Cdc4 prevents mislocalization and CIN in unperturbed cells.

Rational In Silico Design of Molecularly Imprinted Polymers: Current Challenges and Future Potential.

The rational design of molecularly imprinted polymers has evolved along with state-of-the-art experimental imprinting strategies taking advantage of sophisticated computational tools. In silico methods enable the screening and simulation of innovative polymerization components and conditions superseding conventional formulations. The combined use of quantum mechanics, molecular mechanics, and molecular dynamics strategies allows for macromolecular modelling to study the systematic translation from the pre- to the post-polymerization stage. However, predictive design and high-performance computing to advance MIP development are neither fully explored nor practiced comprehensively on a routine basis to date. In this review, we focus on different steps along the molecular imprinting process and discuss appropriate computational methods that may assist in optimizing the associated experimental strategies. We discuss the potential, challenges, and limitations of computational approaches including ML/AI and present perspectives that may guide next-generation rational MIP design for accelerating the discovery of innovative molecularly templated materials.

The Indian REgistry on Current Patient PrOfiles and TReatment TrenDs in Hypertension (RECORD): Final Outcomes of the Real-world Observational Study.

Objectives: The Indian Registry on Current Patient Profiles and Treatment Trends in Hypertension (Record) evaluated the current trends and outcomes related to hypertension (HTN) management at 3, 6, 12, and 24 months in India. This study highlights and evaluates the outcomes and trends noted at 24 months. Materials and methods: The detailed study methodology is provided in the earlier publication (interim analysis at 12 months). Aspects such as changes in the quality of life (QOL), percentage of patients reaching target blood pressure (BP), treatment pattern among patients with comorbid conditions, and difference in treatment patterns between public and private healthcare settings, at 24 months, were evaluated in the current study. Results: The study population included 2,000 patients (55.7% males) with a mean age of 54.45 years. Telmisartan (43.7%) and amlodipine + telmisartan (16.4%) were the most prescribed monotherapy and combination therapy among patients with newly diagnosed HTN. A significant decrease in both systolic BP (SBP) and diastolic BP (DBP) was noted in the overall patient population at 24 months (p < 0.001). The mean change in SBP and DBP was slightly higher at 24 months compared to 12 months. This was more evident among patients on combination therapy. A significant improvement in QOL was noted at 24 months. Conclusion: Treatment strategies in HTN management are changing and are associated with effective HTN control and improvements in QOL. However, there is a further need for improved awareness regarding the optimal usage of combination therapy for better management of uncontrolled HTN. How to cite this article: Rajadhyaksha GC, Reddy H, Singh AK, et al. The Indian REgistry on Current Patient PrOfiles and TReatment TrenDs in Hypertension (RECORD): Final Outcomes of the Real-World Observational Study. J Assoc Physicians India 2023;71(11):43-49.

A comparison of caries experience among orphanage children with non-orphans attending government school of indore city.

Introduction: Oral health is an imperative to general health. It is important in many aspects of child development, as poor oral health can lead to problems with nutrition, speech development and self-esteem. Children living in orphanage are considered vulnerable to oral diseases. Objective: To identify and compare the caries experience of children between the ages of 6 and 15 living in orphanages with children attending school in the city of Indore. Methods: A descriptive cross-sectional study was conducted among 6-15 years aged orphanage children and children studying in schools located in the same geographical area of the Indore city. A total of 200 children in each group were taken under the study. The data collected were oral hygiene practice and dentition status on WHO form 2013 for adults. The data was then analysed to determine mean DMFT and deft score. Results: A statistically significant (p=0.001) difference in mean DMFT between orphans and non-orphans was observed. The decayed and missing component shows a statistically significant (p=0.001) difference between the orphans and non-orphans. For the primary dentition, the results show that the mean deft of orphans (0.28±0.84) was significantly higher (p=0.001) than non-orphans. Conclusions: Based on the results of the present study, it can be concluded that the dental caries experience of orphans living in government-funded orphanage homes was found to be better than non-orphans studying in government school.

Adverse Drug Reaction Patterns of First-line Anti-tubercular Drugs among Saharia Tuberculosis Patients: An Observational Study in Particularly Vulnerable Tribal Group of Madhya Pradesh, India.

SUMMARY: The Saharia tribe of Madhya Pradesh has a very high tuberculosis (TB) burden. However, there is no report of adverse drug reaction (ADR) available in patients receiving anti-TB chemotherapy in the community. Reporting and monitoring of ADRs among TB patients is still rare in marginalized communities. An observational prospective study was performed from November 2019 to June 2020 to assess the patterns of ADRs in 250 Saharia TB patients, who were prescribed Category-I daily DOTS (HRZE) by the physician. Both male and female participants equally experienced ADR during the treatment, but relatively more females (92.6%) than males (88.6%) reported ADR during Phase I. Out of 250 patients, 224 patients (89.6%) experienced one or more ADRs in Phase I. The central nervous system-related (75.6%) ADR was mostly reported followed by any gastrointestinal (74.4%), cardiovascular (49.2%) and any dermatological related (44.4%) ADRs. It is paramount to timely monitor and proactively manages ADRs pertaining to anti-TB drug treatment with minimal alteration in the treatment course.

Evidence-based mechanistic role of chrysin towards protection of cardiac hypertrophy and fibrosis in rats.

Cardiac hypertrophy is the enlargement of cardiomyocytes in response to persistent release of catecholamine which further leads to cardiac fibrosis. Chrysin, flavonoid from honey, is well known for its multifarious properties like antioxidant, anti-inflammatory, anti-fibrotic and anti-apoptotic. To investigate the cardioprotective potential of chrysin against isoproterenol (ISO), cardiac hypertrophy and fibrosis are induced in rats. Acclimatised male albino Wistar rats were divided into seven groups (n 6): normal (carboxymethyl cellulose at 0·5 % p.o.; as vehicle), hypertrophy control (ISO 3 mg/kg, s.c.), CHY15 + H, CHY30 + H & CHY60 + H (chrysin; p.o.15, 30 and 60 mg/kg respectively + ISO at 3 mg/kg, s.c.), CHY60 (chrysin 60 mg/kg in per se) and LST + H (losartan 10 mg/kg p.o. + ISO 3 mg/kg, s.c.) were treated for 28 d. After the dosing schedule on day 29, haemodynamic parameters were recorded, after that blood and heart were excised for biochemical, histological, ultra-structural and molecular evaluations. ISO administration significantly increases heart weight:body weight ratio, pro-oxidants, inflammatory and cardiac injury markers. Further, histopathological, ultra-structural and molecular studies confirmed deteriorative changes due to ISO administration. Pre-treatment with chrysin of 60 mg/kg reversed the ISO-induced damage to myocardium and prevent cardiac hypertrophy and fibrosis through various anti-inflammatory, anti-apoptotic, antioxidant and anti-fibrotic pathways. Data demonstrated that chrysin attenuated myocardial hypertrophy and prevented fibrosis via activation of transforming growth factor-beta (TGF-ß)/Smad signalling pathway.

The ubiquitin ligase UBE3B, disrupted in intellectual disability and absent speech, regulates metabolic pathways by targeting BCKDK.

Kaufman oculocerebrofacial syndrome (KOS) is a recessive neurodevelopmental disorder characterized by intellectual disability and lack of speech. KOS is caused by inactivating mutations in UBE3B, but the underlying biological mechanisms are completely unknown. We found that loss of Ube3b in mice resulted in growth retardation, decreased grip strength, and loss of vocalization. The brains of Ube3b-/- mice had hypoplasia of the corpus callosum, enlarged ventricles, and decreased thickness of the somatosensory cortex. Ube3b-/- cortical neurons had abnormal dendritic morphology and synapses. We identified 22 UBE3B interactors and found that branched-chain α-ketoacid dehydrogenase kinase (BCKDK) is an in vivo UBE3B substrate. Since BCKDK targets several metabolic pathways, we profiled plasma and cortical metabolomes from Ube3b-/- mice. Nucleotide metabolism and the tricarboxylic acid cycle were among the pathways perturbed. Substrate-induced mitochondrial respiration was reduced in skeletal muscle but not in liver of Ube3b-/- mice. To assess the relevance of these findings to humans, we identified three KOS patients who had compound heterozygous UBE3B mutations. We discovered changes in metabolites from similar pathways in plasma from these patients. Collectively, our results implicate a disease mechanism in KOS, suggest that it is a metabolic encephalomyopathy, and provide an entry to targeted therapies.

Evaluation of Crestal Bone Loss in Flap vs Flapless Implant Placement Technique: An In Vivo Comparative Study.

AIMS: The aims of the study were to assess the technique with-flap and flapless implant placement and to compare crestal bone heights around the implant in flapless and conventional flap technique using digital radiovisiograph, in 3 and 6 months after the surgery. MATERIALS AND METHODS: A total of 20 implants were placed by flap and flapless implant technique; each patient received two implants, except for two patients who received four implants. A radiovisiograph was taken at implant placement, as well as 3- and 6-month intervals. Crestal bone level was compared between flapless and flap during these intervals and compared between intervals for each group. RESULTS: On evaluating the distribution, it was found to be asymmetric and hence lacked normality (K-S = 0.382; p <0.001). On mesial side, bone loss values in group I ranged from 0.40 to 1.10 units with a mean value of 0.71 and a standard deviation of 0.26 units. The median value was 0.70. On evaluating the data for normality, it was found to be symmetric and normal (K-S = 0.166; p = 0.200). CONCLUSION: This study concluded that there are not any significant differences in the crestal bone with both flap and flapless techniques. Comparatively, the flapless approach showed a lesser crestal bone height reduction, which was statistically significant. CLINICAL SIGNIFICANCE: Implant dentistry is nonetheless behind when advances are concerned, we have seen the inclination toward minimal invasive implant techniques to yield better esthetic as well as improved results, thus taking care of patients' discomfort.

Extensively drug-resistant tuberculosis treated with bedaquiline: A case report in the particularly vulnerable tribal group of Madhya Pradesh, India.

The management of drug-resistant (DR) tuberculosis (TB) remains a challenge particularly in remote rural areas of the country. Although the treatment with wholly oral drug regimens, including bedaquiline (BDQ) and delamanid, is rolled out under the National TB Elimination Program, little is known about its coverage and the effectiveness in hard-to-reach tribal areas. The present report describes the early identification and successful management, through team effort, of a case of extensively DR TB belonging to the Saharia tribe - a Particularly Vulnerable Tribal Groups (PVTGs) of Madhya Pradesh, which has a very high prevalence of TB. The BDQ-containing regimen was well tolerated and found effective with minimal side effects and contributed to the reduced time to culture conversion and radiological improvements. The concerted efforts and strategies need to be adopted for effective implementation of Programmatic management of DR TB (PMDT) guidelines in remote tribal areas of the country.

α-ketobutyrate links alterations in cystine metabolism to glucose oxidation in mtDNA mutant cells.

Pathogenic mutations in the mitochondrial genome (mtDNA) impair organellar ATP production, requiring mutant cells to activate metabolic adaptations for survival. Understanding how metabolism adapts to clinically relevant mtDNA mutations may provide insight into cellular strategies for metabolic flexibility. In this study, we use 13C isotope tracing and metabolic flux analysis to investigate central carbon and amino acid metabolic reprogramming in isogenic cells containing mtDNA mutations. We identify alterations in glutamine and cystine transport which indirectly regulate mitochondrial metabolism and electron transport chain function. Metabolism of cystine can promote glucose oxidation through the transsulfuration pathway and the production of α-ketobutyrate. Intriguingly, activating or inhibiting α-ketobutyrate production is sufficient to modulate both glucose oxidation and mitochondrial respiration in mtDNA mutant cells. Thus, cystine-stimulated transsulfuration serves as an adaptive mechanism linking glucose oxidation and amino acid metabolism in the setting of mtDNA mutations.

Effect of rutin on pharmacokinetic modulation of diclofenac in rats.

Diclofenac is an extensively used nonsteroidal anti-inflammatory drug, but gastrointestinal liabilities and cardiovascular complications take the shine away from such a widely prescribed drug. On the other hand, rutin, a dietary bioflavonoid, has quite a few pharmacological attributes to improve the efficacy and reduce the dose-related toxicities of diclofenac through the intended food-drug/herb-drug interaction. The aim of the present research work was to investigate the role of rutin on pharmacokinetic modulation and the consequent efficacy of diclofenac. At first, pharmacodynamics and pharmacokinetics of diclofenac as alone and in the presence of rutin were investigated orally in a rat model. Then, mechanistic studies were performed to explain the effect of rutin on improvement in oral exposure as well as the efficacy of diclofenac using a battery of in-vitro/in-situ/in-vivo studies. Results displayed that rutin enhanced efficacy as well as oral bioavailability of diclofenac in rats. A marked increase in permeability of diclofenac by rutin was displayed that is linked to inhibition of Breast Cancer Resistance Protein (BCRP) transporters. There was no significant effect of rutin on the modulation of intestinal transit, CYP2C9 inhibition in human liver microsomes, and CYP2C9/CYP2C11 expression in rat liver tissues to boost the oral exposure of diclofenac. Rutin is found to be an inhibitor for BCRP transporters and can act as an oral bioavailability enhancer for a drug like diclofenac.

Protein kinases in mitotic phosphorylation of budding yeast CENP-A.

Centromere identity is specified epigenetically by specialized nucleosomes containing the evolutionarily conserved centromeric histone H3 variant (Cse4 in budding yeast, CENP-A in humans) which is essential for faithful chromosome segregation. However, the mechanisms of epigenetic regulation of Cse4 have not been clearly defined. We have identified two kinases, Cdc5 (Plk1 in humans) and Ipl1 (Aurora B kinase in humans) that phosphorylate Cse4 to prevent chromosomal instability (CIN). Cdc5 associates with Cse4 in mitosis and Cdc5-mediated phosphorylation of Cse4 is coincident with the centromeric enrichment of Cdc5 during metaphase. Defects in Cdc5-mediated Cse4 phosphorylation causes CIN, whereas constitutive association of Cdc5 with Cse4 results in lethality. Cse4 is also a substrate for Ipl1 and phospho-mimetic cse4 mutants suppress growth defects of ipl1 and Ipl1 kinetochore substrate mutants, namely dam1 spc34 and ndc80. Ipl1-mediated phosphorylation of Cse4 regulates kinetochore-microtubule interactions and chromosome biorientation. We propose that collaboration of Cdc5- and Ipl1-mediated phosphorylation of Cse4 modulates kinetochore structure and function, and chromosome biorientation. These findings demonstrate how phosphorylation of Cse4 regulates the integrity of the kinetochore, and acts as an epigenetic marker for mitotic control.