Oil/water (O/W) nanoemulsions developed from essential oil extracted from wildly growing Calotropis gigantea (Linn.) Aiton F.: synthesis, characterization, stability and evaluation of anti-cancerous, anti-oxidant, anti-inflammatory and anti-diabetic activities.

Calotropis gigantea essential oil is utilized in outmoded medicine, therapeutics, and the cosmetic industries. However, the extreme volatility, oxidation susceptibility, and instability of this oil restricts its application. Thus, encapsulation is a more effective method of shielding this oil from unfavorable circumstances. The creation of oil/water (O/W) nanoemulsions based on Calotropis gigantea essential oil (CEO), known as CNE (Calotropis gigantea essential oil nanoemulsions), and an assessment of its biological potential were the goals of this work. UV, fluorescence, and FT-IR methods were used for physiological characterization. Biological activities, including anti-inflammatory, anti-diabetic, and anti-cancer effects. Studies on the pharmacokinetics of CNE were conducted. CNEs encapsulation efficiency was found to be 92%. The CNE nanoemulsions had a spherical shape with polydispersity index of 0.531, size of 200 nm, and a zeta potential of -35.9 mV. Even after being stored at various temperatures for 50 days, CNE nanoemulsions remained stable. Numerous tests were used to determine the antioxidant capacity of CNE, and the following IC50 values (µl/mL) were found: iron chelating assay: 18, hydroxyl radical scavenging: 37, and nitric oxide radical scavenging activity: 58. The percentage of HeLa cells that remained viable after being treated with CNE was 41% at a higher dose of 1 µl. CNE inhibited α-amylase in a dose-dependent manner, with 72% inhibition at its higher dose of 250 µL. Research on the kinetics of drugs showed that nanoemulsions showed Higuchi pattern. This research showed potential use of Calotropis gigantea oil-based nanoemulsions in the food, cosmetic, and pharmaceutical industries.

Plasmodium falciparum cysteine protease Falcipain 3: A potential enzyme for proteolytic processing of histone acetyltransferase PfGCN5.

In spite of 150 years of studying malaria, the unique features of the malarial parasite, Plasmodium, still perplex researchers. One of the methods by which the parasite manages its gene expression is epigenetic regulation, the champion of which is PfGCN5, an essential enzyme responsible for acetylating histone proteins. PfGCN5 is a ∼170 kDa chromatin-remodeling enzyme that harbors the conserved bromodomain and acetyltransferase domain situated in its C-terminus domain. Although the PfGCN5 proteolytic processing is essential for its activity, the specific protease involved in this process still remains elusive. Identification of PfGCN5 interacting proteins through immunoprecipitation (IP) followed by LC-tandem mass spectrometry analysis revealed the presence of food vacuolar proteins, such as the cysteine protease Falcipain 3 (FP3), in addition to the typical members of the PfGCN5 complex. The direct interaction between FP3 and PfGCN5 was further validated by in vitro pull-down assay as well as IP assay. Subsequently, use of cysteine protease inhibitor E64d led to the inhibition of protease-specific processing of PfGCN5 with concomitant enrichment and co-localization of PfGCN5 and FP3 around the food vacuole as evidenced by confocal microscopy as well as electron microscopy. Remarkably, the proteolytic cleavage of the nuclear protein PfGCN5 by food vacuolar protease FP3 is exceptional and atypical in eukaryotic organisms. Targeting the proteolytic processing of GCN5 and the associated protease FP3 could provide a novel approach for drug development aimed at addressing the growing resistance of parasites to current antimalarial drugs.

Tear Protein Markers for Diabetic Retinopathy and Diabetic Macular Edema - Towards an Early Diagnosis and Better Prognosis.

BACKGROUND: Diabetic eye disease is a highly prevalent and sight-threatening disorder. It is a disease of neuro-vascular unit of the retina, if left untreated can cause blindness. Therapeutic approaches followed for its treatment can only restrict the progression of the disease with highly variable results. There is no known biomarker for an early diagonsis of this disease, therefore by the time it is detected it goes beyond repair. This creates a massive demand for development of such biomarkers that help detect disease in its earlier stages. METHODS: PUBMED (https://pubmed.ncbi.nlm.nih.gov/) was searched for articles relevant to the topic published till November 2023. The search was made using keywords such as Diabetic Retinopathy, inflammation, tear, biomarker, proteomics etc. The studies providing relevant information to prove the importance of biomarker discovery were chosen. After compiling the data, the manuscript writing was planned under relevant headings and sub-headings. RESULTS: The review provides a comprehensive overview of all the tear protein biomarker studies in the field of DR and DME. Briefly, their potential in other diseases is also elucidated. While there are many studies pertaining to DR biomarkers, the identified markers lack validations which has restricted their usage in clinics. In case of DME, there was no such study towards biomarker discovery for its diagnosis and prognosis. CONCLUSIONS: The review highlights major studies and their lacunae in the field of biomarkers discovery for DR and DME.

Compositional profiling and molecular docking studies of Eucalyptus polybrachtea essential oil against mucormycosis and aspergillosis.

Essential oil (EO) from Eucalyptus polybrachtea is used as complementary and traditional medicine worldwide. The present study aimed at compositional profiling of EO and molecular docking of EO's bioactive compound 1,8 cineole against fungal enzymes involved in the riboflavin synthesis pathway, namely riboflavin synthase (RS), riboflavin biosynthesis protein RibD domain-containing protein (RibD), and 3,4-dihydroxy-2-butanone 4-phosphate synthase (DBPS) as apposite sites for drug designing against aspergillosis and mucormycosis, and in vitro confirmation. The compositional profile of EO was completed by GC-FID analysis. For molecular docking, the Patchdock tool was used. The ligand-enzyme 3-D interactions were examined, and ADMET properties (absorption, distribution, metabolism, excretion, and toxicity) were calculated. GC-FID discovered the occurrence of 1,8 cineole as a major component in EO, which was subsequently used for docking analysis. The docking analysis revealed that 1,8 cineole actively bound to RS, RibD, and DBPS fungal enzymes. The results of the docking studies demonstrated that the ligand 1,8 cineole exhibited H-bond and hydrophobic interactions with RS, RibD, and DBPS fungal enzymes. 1,8 cineole obeyed Lpinsky's rule and exhibited adequate bioactivity. Wet-lab authentication was achieved by using three fungal strains: Aspergillus niger, Aspergillus oryzae, and Mucor sp. Wet lab results indicated that EO was able to inhibit fungal growth.

Nanoemulsions (O/W) prepared from essential oil extracted from Melaleuca alternifolia: synthesis, characterization, stability and evaluation of anticancerous, anti-oxidant, anti-inflammatory and antidiabetic activities.

Essential oil from Melaleuca alternifolia (also known as Tea tree essential oil, TTO) is used as traditional medicine and used as therapeutic in medicine, food and cosmetic sectors. However, this oil is highly unstable, volatile and prone to oxidation which limits its practical use. The objective of this study was synthesis of tea tree oil based O/W (oil/water) nanoemulsions (tea tree essential oil nanoemulsions, TNE) and evaluation of its biological potential. Physiological characterization was carried out using UV, fluorescent, and FT-IR techniques. Various biological activities such as anticancerous, antidiabetic and anti-inflammatory were also estimated. Pharmacokinetics study on TNE was carried out. Encapsulation efficiency of nanoemulsions was found to be 83%. Nanoemulsions were spherical in shape with globule size 308 nm, zeta potential -9.42 and polydispersity index was 0.31. Nanoemulsions were stable even after 50 days of storage at different temperatures. Anti-oxidant potential of TNE was conducted by various assays and IC50 were: Nitric oxide radical scavenging activity:225.1, DPPH radical scavenging activity:30.66, Iron chelating assay:38.73, and Iron reducing assay:39.36. Notable anticancer activity was observed with the percent cell viability of HeLa cells after treatment with 1, 2 and 5 µl of TNE was 82%, 41% and 24%, respectively. Antidiabetic study revealed that TNE inhibited -amylase in a dose-dependent manner, with 88% inhibition at its higher volume of 250 µl. Drug kinetic study revealed that nanoemulsions exhibited first-order model. Based on this, the possible role of M. alternifolia oil-based nanoemulsions in cosmetic, food, and pharma sectors has been discussed.

Targeting collagen XVIII improves the efficiency of ErbB inhibitors in breast cancer models.

The tumor extracellular matrix (ECM) critically regulates cancer progression and treatment response. Expression of the basement membrane component collagen XVIII (ColXVIII) is induced in solid tumors, but its involvement in tumorigenesis has remained elusive. We show here that ColXVIII was markedly upregulated in human breast cancer (BC) and was closely associated with a poor prognosis in high-grade BCs. We discovered a role for ColXVIII as a modulator of epidermal growth factor receptor tyrosine kinase (ErbB) signaling and show that it forms a complex with ErbB1 and -2 (also known as EGFR and human epidermal growth factor receptor 2 [HER2]) and α6-integrin to promote cancer cell proliferation in a pathway involving its N-terminal portion and the MAPK/ERK1/2 and PI3K/AKT cascades. Studies using Col18a1 mouse models crossed with the mouse mammary tumor virus-polyoma virus middle T antigen (MMTV-PyMT) mammary carcinogenesis model showed that ColXVIII promoted BC growth and metastasis in a tumor cell-autonomous manner. Moreover, the number of mammary cancer stem cells was significantly reduced in the MMTV-PyMT and human cell models upon ColXVIII inhibition. Finally, ablation of ColXVIII substantially improved the efficacy of ErbB-targeting therapies in both preclinical models. In summary, ColXVIII was found to sustain the stemness properties of BC cells and tumor progression and metastasis through ErbB signaling, suggesting that targeting ColXVIII in the tumor milieu may have important therapeutic potential.

Underutilized Plant Cymbopogan martinii Derived Essential Oil Is Excellent Source of Bioactives with Diverse Biological Activities.

Cymbopogan martinii, also known as Palmarosa, is an underutilized plant of tropical region. Due to outstanding antioxidant potential it has been used as a part of conventional medicine and beauty product. Regardless of its importance, complete pharmacological and phytochemical studies are still in its early stages. In the current study, Palmarosa essential oil (PRO) was extracted from Cymbopogan martinii and was evaluated for its phytochemicals, antimicrobial and antifungal, anti-inflammatory and anti-diabetic and protection from UV rays. Oil from fresh leaves was extracted and analysed for presence of phytochemicals (Tannin, Flavonoids, and Phenolics). Various antioxidant activities like DPPH (1,1-diphenyl-2-picrylhydrazyl), ABTS (2,2-azinobis-3-ethylbenzothiazoline-6-sulphonic acid), Nitric oxide radical, Hydroxyl radical, iron reducing, iron cheating activity were performed. Antibacterial, anti-inflammatory, Antidiabetic, membrane integrity assay, and UV-absorption assay was also performed. Antifungal activity against "Aspergillosis" and "Mucormycosis" causing fungal strains was also evaluated. High concentration of polyphenolics like Tannin, Flavonoid, phenolics were revealed through phytochemical analysis. GC-FID revealed the presence of Geraniol, major component of Palmarosa oil and other bioactive compound in PRO. PRO showed high anti-inflammatory and anti-diabetic potential and can be used as an Antidiabetic agent due to inhibitory effect on α-amylase activity. Further study revealed that PRO inhibits α-amylase in competitive manner. Hence from the results obtained it is confirmed that the PRO possesses considerable amount of bioactive compounds and can be used in pharmaceutical, food and cosmetic industries.

Essential Oil Derived from Underutilized Plants Cymbopogon khasianus Poses Diverse Biological Activities against "Aspergillosis" and "Mucormycosis".

Palmrosa essential oil (PEO) from Cymbopogon khasianus, is used as complementary and traditional medicine worldwide. The present study aimed at compositional profiling of PEO and molecular docking of PEO bioactive compound geraniol against fungal enzymes chitin synthase (CS), UDP-glycosyltransferase (UDPG) and glucosamine-6-phosphate synthase (GPS), as apposite sites for drug designing against "Aspergillosis" and "Mucormycosis" and in vitro confirmation. Compositional profile of PEO was completed by GC-FID analysis. For molecular docking, Patch-dock tool was conducted. Ligand-enzyme 3D interactions were also calculated. ADMET properties (absorption, distribution, metabolism, excretion and toxicity) were also calculated. GC-FID discovered the occurrence of geraniol as a major component in PEO, thus nominated for docking analysis. Docking analysis specified active binding of geraniol to GPS, CS and UDPG fungal enzymes. Wet-lab authentication was achieved by three fungal strains Aspergillus niger, A. oryzae and Mucor sp. Docking studies revealed that ligand geraniol exhibited intercations with GPS, CS and UDPG fungal enzymes by H-bond and hydrophobic interactions. Geraniol obeyed LIPINSKY rule, and exhibited adequate bioactivity. Wet lab results indicated that PEO was able to inhibit fungal growth against "Aspergillosis" and "Mucormycosis".

Combining in silico and in vitro approaches to understand the involvement of methylerythritol 4-phosphate and shikimate pathways in Agrobacterium tumefaciens for enhanced coenzyme Q10 production.

AIMS: To perform an integrated comparative analysis of metabolic pathway to understand coenzyme Q10 (CoQ10) production in Agrobacterium tumefaciens. METHODS AND RESULTS: Comparative analysis of the CoQ10 metabolic pathway in 10 organisms using a genome to KEGG orthology program (G2KO) and the KEGG database elucidated the completeness of the production pathway in A. tumefaciens. The specific roles of the key precursors and the enzymes in the metabolic network were subsequently confirmed using pathway inhibitors and enhancers. While the use of fosmidomycin and glyphosate was found to inhibit CoQ10 production by 54.54% to 99%, the supplementation of polyprenyl pyrophosphate of the methylerythritol 4-phosphate pathway and 4-hydroxybenzoate precursor of the shikimate pathway did increse the production of CoQ10 by 2.3-fold. CONCLUSIONS: The present study provides a comprehensive understanding of the CoQ10 biosynthetic pathway in A. tumefaciens, which would assist rational metabolic engineering strategies for augmenting CoQ10 biosynthesis.

Electrospun fiber-based strategies for controlling early innate immune cell responses: Towards immunomodulatory mesh designs that facilitate robust tissue repair.

Electrospun fibrous meshes are widely used for tissue repair due to their ability to guide a host of cell responses including phenotypic differentiation and tissue maturation. A critical factor determining the eventual biological outcomes of mesh-based regeneration strategies is the early innate immune response following implantation. The natural healing process involves a sequence of tightly regulated, temporally varying and delicately balanced pro-/anti-inflammatory events which together promote mesh integration with host tissue. Matrix designs that do not account for the immune milieu can result in dysregulation, chronic inflammation and fibrous capsule formation, thus obliterating potential therapeutic outcomes. In this review, we provide systematic insights into the effects of specific fiber/mesh properties and mechanical stimulation on the responses of early innate immune modulators viz., neutrophils, monocytes and macrophages. We identify matrix characteristics that promote anti-inflammatory immune phenotypes, and we correlate such responses with pro-regenerative in vivo outcomes. We also discuss recent advances in 3D fabrication technologies, bioactive functionalization approaches and biomimetic/bioinspired immunomodulatory mesh design strategies for tissue repair and wound healing. The mechanobiological insights and immunoregulatory strategies discussed herein can help improve the translational outcomes of fiber-based regeneration. STATEMENT OF SIGNIFICANCE: The crucial role played by immune cells in promoting biomaterial-based tissue regeneration is being increasingly recognized. In this review focusing on the interactions of innate immune cells with electrospun fibrous meshes, we systematically elucidate the effects of the fiber microenvironment and mechanical stimulation on biological responses, and build upon these insights to inform the rational design of immunomodulatory meshes for effective tissue repair. We discuss state-of-the-art fabrication methods and mechanobiological advances that permit the orchestration of temporally controlled phenotypic switches in immune cells during different phases of healing. The design strategies discussed herein can also be leveraged to target several complex autoimmune and inflammatory diseases.

Nanoemulsions (O/W) containing Cymbopogon pendulus essential oil: development, characterization, stability study, and evaluation of in vitro anti-bacterial, anti-inflammatory, anti-diabetic activities.

Essential oil from Cymbopogon pendulus is immensely useful in various sectors like food, pharmaceutical, and cosmetic industries. Since this oil is hydrophobic, unstable, and volatile, hence encapsulation by using nanoemulsions technology is the best way to protect it. This study reports biosynthesis of O/W (oil/water) nanoemulsions based on essential oil from Cymbopogon pendulus and analysis of its biological activities. O/W nanoemulsions were prepared by using tween 20/80, sodium dodecyl sulphate as surfactants, and ethanol as co-surfactants. Fingerprinting of nanoemulsions using UV, fluorescent, and FT-IR was studied along with other parameters like pH and conductivity. Biological activities like antibacterial, anti-inflammatory, and anti-diabetic activities and drug release pharmokinetics were evaluated. Ethanol containing nanoemulsions was noticeably smaller than other nanoemulsions. Encapsulation efficiency of nanoemulsions was in the range from 41 to 60%. Nanoemulsions were spherical in shape and stable even after 50 days of storage. Appreciable biological activities like anti-bacterial, anti-inflammatory, and anti-diabetic activities were detected. Drug kinetic study revealed that nanoemulsions exhibited Korsmeyer-Peppas model. Based on this, the possible role of lemon grass oil-based nanoemulsions in cosmetic, food, and pharma sectors has been discussed. Supplementary Information: The online version contains supplementary material available at 10.1007/s12668-022-00964-4.

Essential oil from Cymbopogon citratus exhibits "anti-aspergillosis" potential: in-silico molecular docking and in vitro studies.

BACKGROUND: Aspergillosis, has recently confounded some states of India. Due to major role in fungal cell wall synthesis, in the present study UDP-glycosyltransferase, Glucosamine-6-phosphate synthase and chitin synthase were chosen as an appropriate sites to design drug. The objective of present study was molecular docking of lemon grass essential oil component citral and in vitro validation. GC-FID analysis was used to find out aromatic profile. For docking, Patch-dock analysis was used. Ligand Protein 2D and 3D Interactions were also studied. Drug likeliness, and toxicity profile were also studied. Docking analysis indicated effective binding of citral to UDP-glycosyltransferase, Glucosamine-6-phosphate synthase and chitin synthase. In vitro validation was performed by fungal strain Aspergillus fumigatum. RESULTS: GC-FID profiling revealed the presence of citral as major bioactive compound. Interactions results indicated that, UDP-glycosyltransferase, Glucosamine-6-phosphate synthase and chitin synthase enzymes and citral complexes forms hydrogen and hydrophobic interactions. Citral also depicted drug likeliness by LIPINSKY rule, sufficient level of bioactivity, drug likeliness and toxicity. CONCLUSION: In vitro results revealed that lemon grass oil was able to inhibit growth of fungal strains toxicity thus signifying its role as potent anti-fungal drug. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1186/s42269-022-00711-5.

Congenital central corneal dermoid: A rare entity.

BACKGROUND: To understand the pathogenesis of a central corneal dermoid (CD) in a 12-day-old child, a comparison of CD specimen was done with limbal dermoid (LD) and cadaveric corneal (CC) specimens by immuno-histochemical staining. METHODS: The child underwent penetrating keratoplasty for visual rehabilitation. The corneal tissue was sent for histo-pathological and immunochemistry evaluation. The corneal specimen obtained was compared the origin of central CD with LD and CC based on their antigenic expression profile. RESULTS: Clinically over a period of 75 months post operatively the child maintained a clear graft. Hematoxylin and Eosin staining of LD had a typical morphology including stratified keratinized epithelium, hair shaft with pilo sebaceous glands, eccrine sweat glands, lymphocytes, and blood vessels. Immuno-histochemical staining showed positive stain for Cytokeratin 3 epithelial marker in the epithelium of CC, LD, and CD. Smooth muscle maker (SMA) was identified in LD and CD but not in the CC as it is devoid of blood vessels. Limbal stem cell maker (P63) was detected only in LD. Vimentin, a mesenchymal stem cell marker stained positively in all three tissues of CC, LD, and CD. CONCLUSIONS: Corneal dermoid showed positive staining for mesodermal tissue components compared to both ectodermal and mesodermal components in limbal dermoid suggesting possibly a different origin of corneal dermoid.

Comparative phytochemistry, antioxidant, antidiabetic, and anti-inflammatory activities of traditionally used Ocimum basilicum L. Ocimum gratissimum L., and Ocimum tenuiflorum L.

Ocimum spp. are the widely studied herbal plants because of their diverse biological activities. The present study aimed at comparative extraction of secondary metabolites and evaluation of their biological activities in different solvents such as acetone, ethanol, methanol, and water. Three Ocimum species, namely Ocimum basilicum L. (Green tulsi), Ocimum gratissimum L. (Jungli tulsi), and Ocimum tenuiflorum (Black tulsi), were selected for this study. Leaf extracts from dried powder of these species were prepared in different solvents. The contents of total phenolics, flavonoids, and total condensed tannins were estimated using standard assays. Fingerprint analysis using UV, Fourier transform infrared (FT-IR), and fluorescent spectroscopy was also conducted. Total antioxidant, antidiabetic, and anti-inflammatory activities of the extracts were evaluated. Fingerprint analysis indicated the presence of a sufficient level of polyphenolics in all the solvent extracts. Among all the solvents, acetone provided a higher yield of phenolics, flavonoids, and tannins in all Ocimum species. Black Ocimum showed the maximum level of antioxidants. All Ocimum extracts exhibited a sufficient level of antidiabetic and anti-inflammatory activities. The results indicated that by using appropriate solvents, bioactive compounds from Ocimum species can be extracted and used as therapeutic agents with potential biological activities.

To study the role of pre-treatment microRNA (micro ribonucleic acid) expression as a predictor of response to chemoradiation in locally advanced carcinoma cervix.

BACKGROUND: Concurrent chemoradiotherapy followed by brachytherapy is the standard of care in locally advanced carcinoma cervix. There is no prognostic factor at present to predict the outcome of disease in locally advanced carcinoma cervix. AIM: Differential expression of microRNAs can be used as biomarkers to predict clinical response in locally advanced carcinoma cervix patients. METHODS: Thirty-two patients of locally advanced carcinoma cervix with International Federation of Gynecology and Obstetrics Stage IB-IVA were enrolled from 2017 to 2018. Expression of microRNA-9 5p, -31 3p, -100 5p, -125a 5p, -125b-5p, and -200a 5p in formalin-fixed paraffin embedded (FFPE) biopsied tissue were analyzed by real time quantitative reverse transcriptase polymerase chain reaction (RT qPCR). Pretreatment evaluation was done with clinical examination and MRI pelvis. All patients received concurrent chemoradiotherapy followed by brachytherapy. Patients were evaluated for the clinical response after 3 months of treatment, with clinical examination and MRI pelvis scan using RECIST 1.1 criteria. Patients with no residual disease were classified as Complete responders (CR) and with residual or progressive disease were classified as Nonresponders (NR). Results were statistically analyzed using Mann Whiney U test to examine significant difference between the expression of microRNA between complete responders (CR) and nonresponders (NR). RESULTS: microRNA-100 5p was upregulated in complete responders (CR) which showed a trend towards statistical significance (p value = 0.05). CONCLUSION: microRNA-100 5p can serve as a potential molecular biomarker in predicting clinical response to chemoradiation in locally advanced Carcinoma cervix. Its role should be further investigated in a larger study population.

Management of cervical cancer during the corona virus disease-19 (COVID-19) era.

COVID-19 pandemic has had a catastrophic impact on the society, economy and heath-care system all over the globe with virus showing no signs of losing potency. As the situation appears to worsen, extra burden on other specialities like oncology seems to increase. Specific recommendations are necessary for management of cervical cancer in the current context. All concerned specialities must work together in the best interest of the patient. Attempts should be made at managing cervical cancer while limiting the viral spread among the patients and health-care workers without the loss of opportunity. Surgical intervention for early cervical cancer should be postponed or alternative modalities be considered. In a locally advanced disease, concurrent chemoradiation is the treatment of choice. In addition, the following under mentioned suggestions aim to discuss ways of minimizing infection spread, workload rationalization and providing guidance for management of cervical cancer in the presence of COVID-19 infection.

Antibacterial action of acriflavine hydrochloride for eradication of the gastric pathogen Helicobacter pylori.

Helicobacter pylori, a type 1 carcinogen, accounts for numerous gastric cancer-related deaths worldwide. Repurposing existing drugs or developing new ones for a combinatorial approach against increasing antimicrobial resistance is the need of the hour. This study highlights the efficacy of acriflavine hydrochloride (ACF-HCl) in inhibiting the growth of H. pylori reference strain and antibiotic-resistant clinical isolates at low concentrations. ACF-HCl inhibits H. pylori growth at MIC value 10 times less than that in Escherichia coli, another Gram-negative bacteria. Furthermore, ACF-HCl demonstrates synergistic effect with clarithromycin, a commonly used antibiotic against H. pylori. ACF-HCl treatment also eradicates H. pylori infection in the mice model efficiently. Our in vitro data indicate that bacterial membrane is the prime target. The novel action of ACF-HCl against antibiotic-resistant clinical isolates, synergistic effect with the conventional antibiotic clarithromycin and eradication of H. pylori from infected mice highlight the potential of ACF-HCl as a promising therapeutic agent against H. pylori by itself as well as for combinatorial therapy.

Molecular Assessment of Epiretinal Membrane: Activated Microglia, Oxidative Stress and Inflammation.

Fibrocellular membrane or epiretinal membrane (ERM) forms on the surface of the inner limiting membrane (ILM) in the inner retina and alters the structure and function of the retina. ERM formation is frequently observed in ocular inflammatory conditions, such as proliferative diabetic retinopathy (PDR) and retinal detachment (RD). Although peeling of the ERM is used as a surgical intervention, it can inadvertently distort the retina. Our goal is to design alternative strategies to tackle ERMs. As a first step, we sought to determine the composition of the ERMs by identifying the constituent cell-types and gene expression signature in patient samples. Using ultrastructural microscopy and immunofluorescence analyses, we found activated microglia, astrocytes, and Müller glia in the ERMs from PDR and RD patients. Moreover, oxidative stress and inflammation associated gene expression was significantly higher in the RD and PDR membranes as compared to the macular hole samples, which are not associated with inflammation. We specifically detected differential expression of hypoxia inducible factor 1-α (HIF1-α), proinflammatory cytokines, and Notch, Wnt, and ERK signaling pathway-associated genes in the RD and PDR samples. Taken together, our results provide new information to potentially develop methods to tackle ERM formation.

Multidrug-Resistant Pseudomonas aeruginosa Evokes Differential Inflammatory Responses in Human Microglial and Retinal Pigment Epithelial Cells.

Increasing incidences of multidrug-resistant (MDR) pathogens causing endophthalmitis threaten our ability to treat this condition, and the modulation of inflammatory responses by MDR bacteria is not known. In this study, using human microglia and retinal pigment epithelial (RPE) cells, we compare the inflammatory responses of sensitive (S-PA) and multidrug-resistant (MDR-PA) clinical isolates of Pseudomonas aeruginosa. Infected cells were subjected to qPCR analysis, enzyme-linked immunosorbent assay (ELISA), and immunostaining to assess the expression of inflammatory mediators. Both microglia and RPE cells, challenged with S-PA and MDR-PA, induced a time-dependent expression of inflammatory cytokines. Significant differences were observed in expression levels of Toll-like receptors (TLR) TLR4, TLR5, and TLR9 in microglia cells challenged with MDR-PA vs. S-PA. Similarly, mRNA levels of interleukin (IL)-6, tumor necrosis factor (TNF)-α, Interferon (IFN)-γ, and matrix metalloproteinase (MMP)-9 were also higher in MDR-PA-infected cells. At protein levels, upregulation was observed for IL-10 (p = 0.004), IL-8 (p = 0.0006), IL-1ß (p = 0.02), and Granulocyte-macrophage colony-stimulating factor (GM-CSF) (p = 0.0006) in cells infected MDR-PA versus S-PA in both microglia and RPE cells; however, the response was delayed in RPE cells. Heatmap and STRING analysis highlighted the existence of a cross-talk between the inflammatory and cytokine-mediated signaling pathways. Our study highlights a differential inflammatory response evoked by MDR vs. sensitive pathogens in retinal cells during endophthalmitis.

A Systematic Investigation on Complement Pathway Activation in Diabetic Retinopathy.

The complement system plays a crucial role in retinal homeostasis. While the proteomic analysis of ocular tissues in diabetic retinopathy (DR) has shown the deposition of complement proteins, their exact role in the pathogenesis of DR is yet unclear. We performed a detailed investigation of the role of the complement system by evaluating the levels of major complement proteins including C3, C1q, C4b, Complement Factor B (CFB), and Complement Factor H (CFH) and their activated fragments from both the classical and alternative pathways in vitreous humor and serum samples from proliferative DR (PDR) patients and controls. Further, the expressions of complements and several other key pro- and anti-angiogenic genes in the serum and vitreous humor were analyzed in the blood samples of PDR and non-PDR (NPDR) patients along with controls without diabetes. We also assessed the pro-inflammatory cytokines and matrix metalloproteinases in the vitreous humor samples. There was a significant increase in C3 and its activated fragment C3bα' (110 kDa) along with a corresponding upregulation of CFH in the vitreous of PDR patients, which confirmed the increased activation of the alternative complement pathway in PDR. Likewise, a significant upregulation of angiogenic genes and downregulation of anti-angiogenic genes was seen in PDR and NPDR cases. Increased MMP9 activity and upregulation of inflammatory markers IL8 and sPECAM with a downregulation of anti-inflammatory marker IL-10 in PDR vitreous indicated the possible involvement of microglia in DR pathogenesis. Further, a significantly high C3 deposition in the capillary wall along with thickening of basement membranes and co-localization of CFH expression with CD11b+ve activated microglial cells in diabetic retina suggested microglia as a source of CFH in diabetic retina. The increased CFH levels could be a feedback mechanism for arresting excessive complement activation in DR eyes. A gradual increase of CFH and CD11b expression in retina with early to late changes in epiretinal membranes of DR patients indicated a major role for the alternative complement pathway in disease progression.