Warburg Effect as a Novel Mechanism for Homocysteine-Induced Features of Age-Related Macular Degeneration.

Age-related macular degeneration (AMD) is a major cause of blindness. Recent studies have reported impaired glycolysis in AMD patients with a high lactate/pyruvate ratio. Elevated homocysteine (Hcy) (Hyperhomocysteinemia, HHcy) was observed in several clinical studies, reporting an association between HHcy and AMD. We established the effect of HHcy on barrier function, retinal pigment epithelium (RPE) structure, and induced choroidal neovascularization (CNV) in mice. We hypothesize that HHcy contributes to AMD by inducing a metabolic switch in the mitochondria, in which cells predominantly produce energy by the high rate of glycolysis, or "Warburg", effect. Increased glycolysis results in an increased production of lactate, cellular acidity, activation of angiogenesis, RPE barrier dysfunction, and CNV. Evaluation of cellular energy production under HHcy was assessed by seahorse analysis, immunofluorescence, and western blot experiments. The seahorse analysis evaluated the extracellular acidification rate (ECAR) as indicative of glycolysis. HHcy showed a significant increase in ECAR both in vivo using (Cystathionine ß-synthase) cbs+/- and cbs-/- mice retinas and in vitro (Hcy-treated ARPE-19) compared to wild-type mice and RPE cells. Moreover, HHcy up-regulated glycolytic enzyme (Glucose transporter-1 (GlUT-1), lactate dehydrogenase (LDH), and hexokinase 1 (HK1)) in Hcy-treated ARPE-19 and primary RPE cells isolated from cbs+/+, cbs+/-, and cbs-/- mice retinas. Inhibition of GLUT-1 or blocking of N-methyl-D-aspartate receptors (NMDAR) reduced glycolysis in Hcy-treated RPE and improved albumin leakage and CNV induction in Hcy-injected mice eyes. The current study suggests that HHcy causes a metabolic switch in the RPE cells from mitochondrial respiration to glycolysis during AMD and confirms the involvement of NMDAR in this process. Therefore, targeting Glycolysis or NMDAR could be a novel therapeutic target for AMD.

Homocysteine and Age-Related Central Nervous System Diseases: Role of Inflammation.

Hyperhomocysteinemia (HHcy) is remarkably common among the aging population. The relation between HHcy and the development of neurodegenerative diseases, such as Alzheimer's disease (AD) and eye diseases, and age-related macular degeneration (AMD) and diabetic retinopathy (DR) in elderly people, has been established. Disruption of the blood barrier function of the brain and retina is one of the most important underlying mechanisms associated with HHcy-induced neurodegenerative and retinal disorders. Impairment of the barrier function triggers inflammatory events that worsen disease pathology. Studies have shown that AD patients also suffer from visual impairments. As an extension of the central nervous system, the retina has been suggested as a prominent site of AD pathology. This review highlights inflammation as a possible underlying mechanism of HHcy-induced barrier dysfunction and neurovascular injury in aging diseases accompanied by HHcy, focusing on AD.

Implication of N-Methyl-d-Aspartate Receptor in Homocysteine-Induced Age-Related Macular Degeneration.

Age-related macular degeneration (AMD) is a leading cause of vision loss. Elevated homocysteine (Hcy) (Hyperhomocysteinemia) (HHcy) has been reported in AMD. We previously reported that HHcy induces AMD-like features. This study suggests that N-Methyl-d-aspartate receptor (NMDAR) activation in the retinal pigment epithelium (RPE) is a mechanism for HHcy-induced AMD. Serum Hcy and cystathionine-ß-synthase (CBS) were assessed by ELISA. The involvement of NMDAR in Hcy-induced AMD features was evaluated (1) in vitro using ARPE-19 cells, primary RPE isolated from HHcy mice (CBS), and mouse choroidal endothelial cells (MCEC); (2) in vivo using wild-type mice and mice deficient in RPE NMDAR (NMDARR-/-) with/without Hcy injection. Isolectin-B4, Ki67, HIF-1α, VEGF, NMDAR1, and albumin were assessed by immunofluorescence (IF), Western blot (WB), Optical coherence tomography (OCT), and fluorescein angiography (FA) to evaluate retinal structure, fluorescein leakage, and choroidal neovascularization (CNV). A neovascular AMD patient's serum showed a significant increase in Hcy and a decrease in CBS. Hcy significantly increased HIF-1α, VEGF, and NMDAR in RPE cells, and Ki67 in MCEC. Hcy-injected WT mice showed disrupted retina and CNV. Knocking down RPE NMDAR improved retinal structure and CNV. Our findings underscore the role of RPE NMDAR in Hcy-induced AMD features; thus, NMDAR inhibition could serve as a promising therapeutic target for AMD.

A Prospective Study on Correlation of MERI (Middle Ear Risk Index) Score with Surgical Outcome of Tympano-mastoid Surgery in Patients of CSOM.

Chronic otits media (COM) is surgically treatable disease which includes complete removal of disease and improving hearing via ossicular reconstruction. Therefore, thorough assessment of the disease, ossicles and various factors causing it plays a major role in surgical outcome prediction. MERI (Middle ear risk index) is one such tool being used worldwide. Our aim was to evaluate the surgical outcome of tympanomastoid surgery using MERI and find out correlation between them as well as categorize cases into their severity score in a developing country. Observational prospective study conducted in a tertiary care center. 200 patients were included. After complete history and examination, they were given MERI scores and prediction of surgical outcome was done. Postoperatively it was compared with the real outcome of the surgery. Out of 200 patients, 71.5% had mild, 15.5% had moderate and 13% had severe MERI scores preoperatively. There was a success rate of 88.5% in graft uptake and the mean score of hearing benefit (A-B gain) among patients was 8.75 ± 8.82 dB postoperatively. MERI may be used as a prognostic indicator for predicting surgical outcome. Based on the MERI score, chances of surgical success and hearing benefit can be explained to the patient with certain limitations.

Prevelance of Hearing Loss and Factors Association in Chronic Kidney Disease Patients.

Aim: Chronic kidney disease (CKD) is becoming a great concern, especially in developing countries and SNHL among patients with CKD is considerably higher than general population. So, current study aims evaluation and assessment of degree of hearing loss in chronic kidney disease patients using pure tone audiogram and correlation of hearing loss with its contributing factors. Materials and methods:70 patients with CKD were included in this prospective observational cross sectional study performed in tertiary care hospital. A detailed history; general physical, otological examination, blood investigation was done.CKD staging was done using KDIGO guidelines and hearing assessment using pure tone audiometry. Hearing loss association with CKD and contributing factors were studied. Result: Out of 70 participants, SNHL was seen in twenty one participants. Hearing loss was present in 3 out of 22 participants of CKD-3; 6 out of 12 participants of CKD-4 and 12 out of 15 participants of stage CKD-5. A statistically significant association of hearing loss was found with CKD stage, albuminuria, hemodialysis, hypertension and other factors. Discussion: Since hearing impairment may well have a negative impact on the social function of an affected individual, the use of pure tone audiogram to monitor CKD patients should be considered as a routine procedure. Patients with CKD would benefit from early detection of hearing loss which will allow for early rehabilitative measures to be taken.

Isolation of Primary Mouse Retinal Pigmented Epithelium Cells.

The retinal pigmented epithelium (RPE) layer lies immediately behind the photoreceptors and harbors a complex metabolic system that plays several critical roles in maintaining the photoreceptors' function. Thus, the RPE structure and function are essential to sustain normal vision. This manuscript presents an established protocol for primary mouse RPE cell isolation. RPE isolation is a great tool to investigate the molecular mechanisms underlying RPE pathology in the different mouse models of ocular disorders. Furthermore, RPE isolation can help in comparing primary mouse RPE cells isolated from wild-type and genetically modified mice, as well as testing drugs that can accelerate the development of therapy for visual disorders. The manuscript presents a step-by-step RPE isolation protocol; the entire procedure, from enucleation to seeding, takes approximately 4 hours. The media shouldn't be changed for 5-7 days after seeding, to allow the growth of the isolated cells without disturbance. This process is followed by the characterization of morphology, pigmentation, and specific markers in the cells via immunofluorescence. Cells can be passaged a maximum of three or four times.

A Prospective Correlation Study Between Computerized Tomography of Paranasal Sinuses and Nasal Endoscopy Findings in Patients of Chronic Rhinosinusitis Undergoing Functional Endoscopic Sinus Surgery.

In order to diagnose chronic rhinosinusitis (CRS), diagnostic nasal endoscopy (DNE) and computed tomography (CT) scan both are important investigations. But both have their pros and cons, some findings are seen better in DNE and others in CT. Our study aims to correlate DNE and CT findings. 50 patients with CRS were included in this observational prospective study done at tertiary care hospital. Preoperative each patient underwent DNE and got CT scan followed by scoring using Lund Kennedy and Lund Mackay grading respectively. Functional Endoscopic Sinus Surgery (FESS) was performed and intraoperative findings were correlated with CT scan for each of them. The sensitivity of endoscopy was 93.18% and the specificity was 83.33%. Positive predictive value of DNE was 97.62% and negative predictive value was 62.50%. Most of the endoscopy positive patients of CRS were CT positive. Also, the sensitivity of CT PNS was highest for all groups of sinus disease while specificity was high for posterior group of sinuses (81.82%) and frontal sinus (89.19%). Both DNE and CT scan should be used for planning the management of CRS. DNE tells better about middle meatal secretions, condition of mucosa, polyps. But in situations where due to anatomical variation DNE is difficult, CT scan helps us. CT identifies the extent of disease, the anatomical variants and vital relations of PNS. Overdiagnosis through CT is checked by DNE.

Design, Synthesis, and Molecular Docking Studies of Curcumin Hybrid Conjugates as Potential Therapeutics for Breast Cancer.

Curcumin (CUR) has received great attention over the past two decades due to its anticancer, anti-inflammatory, and antioxidant properties. Similarly, Dichloroacetate (DCA), an pyruvate dehydrogenase kinase 1 (PKD1) inhibitor, has gained huge attention as a potential anticancer drug. However, the clinical utility of these two agents is very limited because of the poor bioavailability and unsolicited side effects, respectively. We have synthesized fusion conjugates of CUR and DCA with an amino acids linker to overcome these limitations by utilizing the molecular hybridization approach. The molecular docking studies showed the potential targets of Curcumin-Modified Conjugates (CMCs) in breast cancer cells. We synthesized six hybrid conjugates named CMC1-6. These six CMC conjugates do not show any significant toxicity in a human normal immortalized mammary epithelial cell line (MCF10A) in vitro and C57BL/6 mice in vivo. However, treatment with CMC1 and CMC2 significantly reduced the growth and clonogenic survival by colony-formation assays in several human breast cancer cells (BC). Treatment by oral gavage of a transgenic mouse BC and metastatic BC tumor-bearing mice with CMC2 significantly reduced tumor growth and metastasis. Overall, our study provides strong evidence that CUR and DCA conjugates have a significant anticancer properties at a sub-micromolar concentration and overcome the clinical limitation of using CUR and DCA as potential anticancer drugs.

RAD51AP1 Loss Attenuates Colorectal Cancer Stem Cell Renewal and Sensitizes to Chemotherapy.

DNA damage, induced by either chemical carcinogens or environmental pollutants, plays an important role in the initiation of colorectal cancer. DNA repair processes, however, are involved in both protecting against cancer formation, and also contributing to cancer development, by ensuring genomic integrity and promoting the efficient DNA repair in tumor cells, respectively. Although DNA repair pathways have been well exploited in the treatment of breast and ovarian cancers, the role of DNA repair processes and their therapeutic efficacy in colorectal cancer is yet to be appreciably explored. To understand the role of DNA repair, especially homologous recombination (HR), in chemical carcinogen-induced colorectal cancer growth, we unraveled the role of RAD51AP1 (RAD51-associated protein 1), a protein involved in HR, in genotoxic carcinogen (azoxymethane, AOM)-induced colorectal cancer. Although AOM treatment alone significantly increased RAD51AP1 expression, the combination of AOM and dextran sulfate sodium (DSS) treatment dramatically increased by several folds. RAD51AP1 expression is found in mouse colonic crypt and proliferating cells. RAD51AP1 expression is significantly increased in majority of human colorectal cancer tissues, including BRAF/KRAS mutant colorectal cancer, and associated with reduced treatment response and poor prognosis. Rad51ap1-deficient mice were protected against AOM/DSS-induced colorectal cancer. These observations were recapitulated in a genetically engineered mouse model of colorectal cancer (ApcMin /+ ). Furthermore, chemotherapy-resistant colorectal cancer is associated with increased RAD51AP1 expression. This phenomenon is associated with reduced cell proliferation and colorectal cancer stem cell (CRCSC) self-renewal. Overall, our studies provide evidence that RAD51AP1 could be a novel diagnostic marker for colorectal cancer and a potential therapeutic target for colorectal cancer prevention and treatment. IMPLICATIONS: This study provides first in vivo evidence that RAD51AP1 plays a critical role in colorectal cancer growth and drug resistance by regulating CRCSC self-renewal.

RAD51AP1 Deficiency Reduces Tumor Growth by Targeting Stem Cell Self-Renewal.

RAD51-associated protein 1 (RAD51AP1) plays an integral role in homologous recombination by activating RAD51 recombinase. Homologous recombination is essential for preserving genome integrity and RAD51AP1 is critical for D-loop formation, a key step in homologous recombination. Although RAD51AP1 is involved in maintaining genomic stability, recent studies have shown that RAD51AP1 expression is significantly upregulated in human cancers. However, the functional role of RAD51AP1 in tumor growth and the underlying molecular mechanism(s) by which RAD51AP1 regulates tumorigenesis have not been fully understood. Here, we use Rad51ap1-knockout mice in genetically engineered mouse models of breast cancer to unravel the role of RAD51AP1 in tumor growth and metastasis. RAD51AP1 gene transcript was increased in both luminal estrogen receptor-positive breast cancer and basal triple-negative breast cancer, which is associated with poor prognosis. Conversely, knockdown of RAD51AP1 (RADP51AP1 KD) in breast cancer cell lines reduced tumor growth. Rad51ap1-deficient mice were protected from oncogene-driven spontaneous mouse mammary tumor growth and associated lung metastasis. In vivo, limiting dilution studies provided evidence that Rad51ap1 plays a critical role in breast cancer stem cell (BCSC) self-renewal. RAD51AP1 KD improved chemotherapy and radiotherapy response by inhibiting BCSC self-renewal and associated pluripotency. Overall, our study provides genetic and biochemical evidences that RAD51AP1 is critical for tumor growth and metastasis by increasing BCSC self-renewal and may serve as a novel target for chemotherapy- and radiotherapy-resistant breast cancer. SIGNIFICANCE: This study provides in vivo evidence that RAD51AP1 plays a critical role in breast cancer growth and metastasis by regulating breast cancer stem cell self-renewal.

Inverted papilloma presenting as unilateral proptosis: A case report with review of literature.

Inverted papilloma is an interesting benign tumour arising from lining epithelium of paranasal sinuses which most commonly involves nasal cavity and paranasal sinuses. However, involvement of orbit and intracranial extension without malignant transformation is very rare. We report a case of extensive inverted papilloma of frontal sinus which primarily presented with proptosis, an uncommon presentation. Ophthalmologic symptoms are rare manifestations of paranasal sinus inverted papilloma without malignant transformation and signify extensive disease with possible intracranial extension.

Giant De Novo Pleomorphic Adenoma of Parapharyngeal Space, Can It Cause Spinal Deformity? A Case Report.

STUDY DESIGN: Case report. INTRODUCTION: De novo giant pleomorphic adenoma is a rare tumor of the parapharyngeal space (PPS). Tumors of the PPS can grow to a large size, compromising the space of the upper aerodigestive tract. However, involvement of the paravertebral region is unexpected. In extremely exceptional circumstances, these tumors can produce spinal deformity. CASE REPORT: A 25-year-old man presented with a longstanding mass in the neck and oral cavity. He had complaints of dysphagia, snoring, and restricted neck movements because of the large size of the tumor. Imaging showed a large mass occupying the PPS extending to the paravertebral region and causing deformity of the cervical spine. Excision was done uneventfully via a minimal access transcervical approach. Residual spinal deformity dealt with postoperative physiotherapy with improvement in lordosis and lateral tilt. No tumor recurrence occurred till 26 months of follow-up. CONCLUSION: Tumors of the PPS can grow to a large size and involve the neck and PPS, even causing spinal deformity. Careful evaluation is required for ascertaining the origin of the tumor and deciding the treatment plan.

Deferiprone: Pan-selective Histone Lysine Demethylase Inhibition Activity and Structure Activity Relationship Study.

Deferiprone (DFP) is a hydroxypyridinone-derived iron chelator currently in clinical use for iron chelation therapy. DFP has also been known to elicit antiproliferative activities, yet the mechanism of this effect has remained elusive. We herein report that DFP chelates the Fe2+ ion at the active sites of selected iron-dependent histone lysine demethylases (KDMs), resulting in pan inhibition of a subfamily of KDMs. Specifically, DFP inhibits the demethylase activities of six KDMs - 2A, 2B, 5C, 6A, 7A and 7B - with low micromolar IC50s while considerably less active or inactive against eleven KDMs - 1A, 3A, 3B, 4A-E, 5A, 5B and 6B. The KDM that is most sensitive to DFP, KDM6A, has an IC50 that is between 7- and 70-fold lower than the iron binding equivalence concentrations at which DFP inhibits ribonucleotide reductase (RNR) activities and/or reduces the labile intracellular zinc ion pool. In breast cancer cell lines, DFP potently inhibits the demethylation of H3K4me3 and H3K27me3, two chromatin posttranslational marks that are subject to removal by several KDM subfamilies which are inhibited by DFP in cell-free assay. These data strongly suggest that DFP derives its anti-proliferative activity largely from the inhibition of a sub-set of KDMs. The docked poses adopted by DFP at the KDM active sites enabled identification of new DFP-based KDM inhibitors which are more cytotoxic to cancer cell lines. We also found that a cohort of these agents inhibited HP1-mediated gene silencing and one lead compound potently inhibited breast tumor growth in murine xenograft models. Overall, this study identified a new chemical scaffold capable of inhibiting KDM enzymes, globally changing histone modification profiles, and with specific anti-tumor activities.

Naso-Septal Tuberculosis Mimicking Dermoid in a 3-Year old Child.

INTRODUCTION: Tuberculosis is a communicable disease caused by mycobacterium tuberculosis that primarily affects the lungs. Primary tuberculosis of the nose in the pediatric age group is rare. The diagnosis of this common entity in the present case was challenging. CASE REPORT: We report the case of a 3-year old girl who presented with a painless swelling over the dorsum of the nose for 7 months. Imaging revealed a mass lesion eroding nasal bones, septum and frontal bone with intracranial extension. Endoscopic examination showed a friable mass in the superior aspect of the nasal septum, extending intracranially. Histopathology confirmed the diagnosis of nasal tuberculosis, and the patient improved on category I anti-tubercular therapy. CONCLUSION: Midline nasal swelling in children needs to be differentiated from congenital nasal swelling. A high index of suspicion is required for correct diagnosis of a patient with nasal tuberculosis. Anti-tubercular therapy is the mainstay of treatment.

Deep neck space abscesses in children below 5 years of age and their complications.

OBJECTIVES: To study the outcomes and complications of deep neck space abscesses in children less than 5 years of age over a period of 15 years. METHODS: A retrospective analysis of children less than 5 years of age with deep neck space abscesses over a 15-year period was conducted at a tertiary care centre in India. Patients were evaluated with respect to the clinical, radiological and laboratory findings. All patients underwent surgical incision and drainage of the abscess and pus cultures were obtained. The incidence of complications was recorded. The collected data was tabulated and statistical analysis was done. RESULTS: A total of 510 children less than 5 years of age were identified who were admitted for deep neck space abscess over a 15-year period. The mean age was 23.6 months. The most common organism isolated in the pus was Staphylococcus aureus (21%). The incidence of methicillin resistant S. aureus (MRSA) was 9%. Complications were reported in 10% patients. CONCLUSION: Paediatric deep neck abscesses can be managed with prompt surgical management and intravenous antibiotics. Children less than 2 years of age, and those with multiple abscesses or retropharyngeal abscess were more prone to complications.

Spontaneous temporomandibular joint herniation: a rare case.

Spontaneous herniation of temporomandibular joint into the external auditory canal through the foramen of Huschke is a very rare condition. We describe a case of spontaneous temporomandibular joint herniation in a 35-year-old male, who presented with otorrhea and aural fullness. The herniation was repaired using collagen mesh. A literature review of all the previous reported cases of spontaneous temporomandibular joint was done to study the presenting clinical features and the method of surgical repair.