An Update on Laboratory-Based Diagnostic Biomarkers for Multiple Sclerosis and Beyond.

BACKGROUND: Multiple sclerosis (MS) is an immune-mediated central nervous system (CNS) inflammatory demyelinating disease in which analysis of clinical presentation, imaging studies, and laboratory tests aid in diagnosis. CONTENT: This review discusses laboratory tests ordered to rule out and rule in MS, such as the traditional measurement of cerebrospinal fluid (CSF) IgG index and oligoclonal bands. Biomarkers discovered in the past 2 decades, such as aquaporin-4 (AQP4) antibodies and myelin oligodendrocyte glycoprotein (MOG) antibodies, have been incorporated into clinical practice in the diagnosis of disorders referred to as MS mimics. The importance of test selection, assay methodology, optimal sample for testing, and diagnostic utility of these biomarkers is reviewed. Other laboratory testing that can aid in the differentiation between MS and these biomarker-defined CNS demyelinating diseases is described. There is a focus on emerging biomarkers such as the use of kappa immunoglobulin free light chain concentration in CSF and kappa CSF index measurement as an alternative to oligoclonal bands which has a potential for an improvement in laboratory workflows. Finally, the role of biomarkers of disease activity and prognosis are discussed, including neurofilament light chain, glial fibrillary acidic protein, and myelin basic protein. Future perspectives with improved laboratory testing tools and discovery of additional biomarkers are provided. SUMMARY: Laboratory testing for demyelinating disorders using CSF and serum are routine practices that can benefit from an update, as novel biomarker-defined entities have reduced the potential for MS misdiagnosis, and CSF/serum biomarkers reinstated in the diagnostic criteria of MS.

HAZ, a novel peptide with broad-spectrum antibacterial activity.

Objective: The growing microbial resistance to antibiotics is a global public concern, which creates serious needs for newer antimicrobial agents. Antimicrobial peptides (AMPs) are increasingly exploited in drug development as therapeutic candidates. Here, we aimed to design and characterize a novel peptide with broad spectrum antimicrobial activity. Methods: Hybridization and sequence modification approaches were used to design the novel peptide, named HAZ, aiming at optimizing the physicochemical parameters involved in antimicrobial activity. Peptide activities were assessed alone or combined with different selected antibiotics against various sensitive and drug-resistant bacterial strains. In addition, the hemolysis and the cytotoxic activities of HAZ peptide were evaluated on human red blood cells and epithelial adenocarcinoma cells (A549), respectively. Results: HAZ peptide was sequentially modified to result in favored physicochemical parameters (helicity 95.24 %, hydrophobic ratio 47 %, and net charge of 8 + ). Functional assessment of HAZ revealed significant antimicrobial activity, with MIC values of 15 - 20 µM against tested bacterial strains. It also exhibited biofilm eradication activity at slightly higher concentrations. HAZ-antibiotics combinations exhibited a synergistic action mode that led to dramatic decrease in the MIC values for both HAZ peptide and the antibiotic. Such efficacy was accompanied with minimal hemolytic toxicity on human erythrocytes. Importantly, HAZ displayed promising anticancer activity against human lung cancer cells. Conclusion: Rationally-designed antimicrobial peptides offer promising alternatives to the current antibiotics for management of infectious diseases. HAZ peptide is a broad-spectrum AMP, and a promising candidate for antimicrobial and anticancer drug development.

Repair Mechanisms in Oxidant-Driven Chronic Inflammatory Disease.

The interplay that governs chronic diseases through pathways specifically associated with chronic inflammation remains undefined. Many metabolic events have been identified during the injury and repair process. Nonetheless, the cellular events that control the pathogenesis of inflammation-induced disease have not been fully characterized. We and others reason that chronic inflammatory diseases associated with a cascade of complex network mediators, such as nitric oxide, arachidonic acid metabolites, cytokines, and reactive oxygen species, play a significant role in the governance of alterations in homeostasis, oxidative stress, and thromboatherosclerosis. In this context, we discuss lipid mediators associated with the maintenance of health, including the specialized proresolving mediators that help drive cellular repair. Emphasis is placed on the pathophysiology of chronic metabolic insults involving both the airways and the cardiovascular system during oxidant-driven inflammatory disease. In this review, we highlight new pathways of inquiry that show promise for the identification of those metabolic targets that can improve therapy for chronic inflammation.

A randomized, double-blind, placebo-controlled crossover study of the effects of levetiracetam on cognition, mood, and balance in healthy older adults.

OBJECTIVE: The cognitive and mood effects of levetiracetam (LEV) in older adults are not known. This study compared the cognitive and mood effects of LEV to placebo in healthy older adults. METHODS: Cognitive, mood, and balance variables were compared between LEV and placebo using a randomized, double-blind, placebo-controlled crossover study with two 5-week treatment periods. Healthy volunteers (n = 20) aged 65-80 (mean age 72.4) received either LEV or placebo in which the LEV target dose was 1,000 mg/day. Volunteers, aged 65-80, were without epilepsy to limit potentially confounding the impact of seizures and/or underlying neuropathology on outcomes. LEV was initiated at 250 mg twice a day for 2 weeks, then increased to 500 mg twice a day for 2 weeks, and then tapered to 250 mg twice a day for 1 week. This was randomized with placebo for the two treatment arms. Measures included standardized neuropsychological, mood, and balance tests yielding 32 variables. Balance was assessed using subjective report (e.g., A-B neurotoxicity scale) and objective data (e.g., Berg Balance Scale). RESULTS: Average LEV serum concentration was 16.9 (standard deviation [SD} 7.7). Repeated-measures analysis of variance (ANOVA) found no differences between LEV and placebo phases for 29 (90.6%) of 32 variables including no change in balance. Performance on LEV was better than placebo on a visual memory (MCG Complex Figure Recall; p = 0.007) and two attention tests (Trail Making Test, Part A, p = 0.009; Stroop Interference, p = 0.004). There was a trend for greater irritability and fatigue (POMS Anger and Fatigue) during the LEV phase (p = 0.029, p = 0.035). Effect-size changes were generally small (Cohen d < 0.5). SIGNIFICANCE: LEV was well tolerated in this elderly population in terms of cognition, mood, and balance. When anticonvulsant medication is indicated for older adults, LEV has pharmacokinetic advantages, and these data indicate no adverse impact on cognition or balance.

Pulmonary Abnormalities in Young, Light-Use Waterpipe (Hookah) Smokers.

RATIONALE: Waterpipes, also called hookahs, are currently used by millions of people worldwide. Despite the increasing use of waterpipe smoking, there is limited data on the health effects of waterpipe smoking and there are no federal regulations regarding its use. OBJECTIVES: To assess the effects of waterpipe smoking on the human lung using clinical and biological parameters in young, light-use waterpipe smokers. METHODS: We assessed young, light-use, waterpipe-only smokers in comparison with lifelong nonsmokers using clinical parameters of cough and sputum scores, lung function, and chest high-resolution computed tomography as well as biological parameters of lung epithelial lining fluid metabolome, small airway epithelial (SAE) cell differential and transcriptome, alveolar macrophage transcriptome, and plasma apoptotic endothelial cell microparticles. MEASUREMENTS AND MAIN RESULTS: Compared with nonsmokers, waterpipe smokers had more cough and sputum as well as a lower lung diffusing capacity, abnormal epithelial lining fluid metabolome profile, increased proportions of SAE secretory and intermediate cells, reduced proportions of SAE ciliated and basal cells, markedly abnormal SAE and alveolar macrophage transcriptomes, and elevated levels of apoptotic endothelial cell microparticles. CONCLUSIONS: Young, light-use, waterpipe-only smokers have a variety of abnormalities in multiple lung-related biological and clinical parameters, suggesting that even limited waterpipe use has broad consequences on human lung biology and health. We suggest that large epidemiological studies should be initiated to investigate the harmful effects of waterpipe smoking.

Smoking-Associated Disordering of the Airway Basal Stem/Progenitor Cell Metabotype.

The airway epithelium is a complex pseudostratified multicellular layer lining the tracheobronchial tree, functioning as the primary defense against inhaled environmental contaminants. The major cell types of the airway epithelium include basal, intermediate columnar, ciliated, and secretory. Basal cells (BCs) are the proliferating stem/progenitor population that differentiate into the other specialized cell types of the airway epithelium during normal turnover and repair. Given that cigarette smoke delivers thousands of xenobiotics and high levels of reactive molecules to the lung epithelial surface, we hypothesized that cigarette smoke broadly perturbs BC metabolism. To test this hypothesis, primary airway BCs were isolated from healthy nonsmokers (n = 11) and healthy smokers (n = 7) and assessed by global metabolic profiling by liquid chromatography-mass spectrometry. The analysis identified 52 significant metabolites in BCs differentially expressed between smokers and nonsmokers (P < 0.05). These changes included metabolites associated with redox pathways, energy production, and inflammatory processes. Notably, BCs from smokers exhibited altered levels of the key enzyme cofactors/substrates nicotinamide adenine dinucleotide, flavin adenine dinucleotide, acetyl coenzyme A, and membrane phospholipid levels. Consistent with the high burden of oxidants in cigarette smoke, glutathione levels were diminished, whereas 3-nitrotyrosine levels were increased, suggesting that protection of airway epithelial cells against oxidative and nitrosative stress is significantly compromised in smoker BCs. It is likely that this altered metabotype is a reflection of, and likely contributes to, the disordered biology of airway BCs consequent to the stress cigarette smoking puts on the airway epithelium.

Risperidone Pellets, Pycnogenol®, and Glucomannan Gummy Formulation for Managing Weight Gain and ADHD in Autistic Children.

Gummy formulations are defined as gradually or slowly released solid oral dosage forms. Risperidone is an atypical antipsychotic medication used to treat schizophrenia and autism-related irritability. This study presents the development of visually appealing, patient-tailored medicated gummies that act as a novel pharmaceutical form of Risperidone for pediatrics. In this study, two gummy bases were used, one containing Glucomannan and the other containing Gelatin as a gelling agent, where these gummy bases were loaded with coated Risperidone pellets with a controlled release layer. The final products were evaluated for their pH, viscosity, content uniformity, drug content, and dissolution profile. Both formulas showed proper rheology and met content and weight uniformity standards. The release rates for F1 and F2 in the acidic media were 25% and 11%, respectively, after 2 h. At the same time, a full-release profile for Risperidone was noticed in both formulae at pH 6.8 where the release lasts for 24 h. It can be concluded that the chewable semi-solid dosages (gummies) filled with coated pellets are suitable for pediatric patients since pediatrics have drug-related problems which can be solved using high gastro-resistance coated pellets, which also shows a proper release profile for the drug.

Predictors of medication regimen complexity and its impact on hemoglobin a1c in type 2 diabetes patients: a retrospective analysis in ambulatory care in Makkah City.

BACKGROUND: Type 2 diabetes mellitus (T2DM) is a widespread chronic disease that poses a significant management challenge due to the complexity of the associated medication regimens, which can have a considerable impact on patient outcomes. OBJECTIVES: Explore the complexity level of diabetes medications among patients with T2DM and to identify the predictors of medication regimen complexity (MRC) and its correlation with hemoglobin A1C (HbA1c) levels. DESIGN: Retrospective, cross-sectional study. SETTING: An ambulatory care setting of a tertiary hospital in Makkah City, Saudi Arabia. PATIENTS AND METHODS: Patients with T2DM referred to the diabetic clinic were identified and assessed for eligibility. The data were collected from patient electronic medical records between October 2022 and September 2023. The MRC Index was used to evaluate the complexity of the patients' medication regimens. MAIN OUTCOMES MEASURES: MRC index scores and HbA1c levels. SAMPLE SIZE: 353 records of patients with T2DM. RESULTS: The analysis revealed that 61.8% (n=218) of patients had high MRC, with the dosing frequency contributing significantly to their MRC (mean=3.9, SD=1.9). Having polypharmacy and longstanding T2DM were predictors of high MRC (odds ratios=4.9 and 2.6, respectively; P≤.01). Additionally, there was an inverse association between the patients' diabetes-specific MRC index scores and their glycemic control (odds ratios=0.2, P<.001). CONCLUSION: The study findings highlight the importance of considering MRC in managing T2DM. Simplifying medication regimens and optimizing medication management strategies can improve patient outcomes. Further research is needed to explore interventions to reduce MRC and enhance diabetes management in this population. LIMITATIONS: Retrospective study design measuring the MRC at a diabetes-specific level.

Characterization of a cellular denitrase activity that reverses nitration of cyclooxygenase.

Protein 3-nitrotyrosine (3-NT) formation is frequently regarded as a simple biomarker of disease, an irreversible posttranslational modification that can disrupt protein structure and function. Nevertheless, evidence that protein 3-NT modifications may be site selective and reversible, thus allowing for physiological regulation of protein activity, has begun to emerge. We have previously reported that cyclooxygenase (COX)-1 undergoes heme-dependent nitration of Tyr(385), an internal and catalytically essential residue. In the present study, we demonstrate that nitrated COX-1 undergoes a rapid reversal of nitration by substrate-selective and biologically regulated denitrase activity. Using nitrated COX-1 as a substrate, denitrase activity was validated and quantified by analytic HPLC with electrochemical detection and determined to be constitutively active in murine and human endothelial cells, macrophages, and a variety of tissue samples. Smooth muscle cells, however, contained little denitrase activity. Further characterizing this denitrase activity, we found that it was inhibited by free 3-NT and may be enhanced by endogenous nitric oxide and exogenously administered carbon monoxide. Finally, we describe a purification protocol that results in significant enrichment of a discrete denitrase-containing fraction, which maintains activity throughout the purification process. These findings reveal that nitrated COX-1 is a substrate for a denitrase in cells and tissues, implying that the reciprocal processes of nitration and denitration may modulate bioactive lipid synthesis in the setting of inflammation. In addition, our data reveal that denitration is a controlled process that may have broad importance for regulating cell signaling events in nitric oxide-generating systems during oxidative/nitrosative stress.

Short-Term and Long-Term Stability of Medial Olivocochlear Reflex in Adults with Typical Hearing.

This study aimed to assess the stability of Medial Olivocochlear Reflex (MOCR) function in typical hearing adults with the use of Contralateral Suppression (CS) of Distortion Product Otoacoustic Emissions (DPOAEs). This study included fifty-three (90 ears) participants between the ages of 18-30. Participants were divided into 3 groups (Group A-daily stability, Group B-short-term stability, and Group C- long-term stability). For each group, 4 measurements (30 × 4 = 120sessions) were taken. Group A measurements were taken daily, Group B measurements were taken weekly and Group C measurements were taken monthly. DPOAEs and Contralateral Suppression of DPOAEs were measured for each group. Analyses indicated that Medial Olivocochlear Reflex (MOCR) measured through contralateral suppression of DPOAE was unstable. This result indicates a DPOAE-based measure of the MOCR was not repeated across time. A great deal has been learned using CS of DPOAEs to study medial efferent activation, but several unresolved methodological issues that could impact the data to produce poor stability across time. Those methodological issues need to be explored and researched in the future.

In Vivo Evaluation of Chitosan-Titanium Dioxide Nanopowder as Wound Dressing Material.

BACKGROUND: Films used for wound healing have many advantages, but should be flexible, robust, adherable and prevent maceration. Both Chitosan (CS) and Titanium dioxide nanopowder (TiO2 NP) have good properties to accelerate wound healing and can be used in preparing films. OBJECTIVE: CS and TiO2 NP are combined to formulate films for wound healing. The physical, thermal, chemical, and mechanical characteristics of these films are to be assessed. The antibacterial activity of the films and their performance on wounded rats will be explored. METHODS: Films made of CS and TiO2 NP were characterized by FTIR, TGA, DSC, XRD, and SEM. The films' mechanical characteristics and antimicrobial activity were tested. Films with acceptable mechanical properties were evaluated on rats. RESULTS: Generally, CS-TiO2 films had higher weight and thickness but lowered flexibility compared to films prepared using CS only. The chosen film showed excellent folding endurance with weight and thickness of around 21.98 mg and 0.16 mm. The surface pH for CS-TiO2 films was acidic, and for the selected film, it was 5.18. CS-TiO2 film was active against all studied bacteria and significantly higher than CS films. The antimicrobial activity of Gram-negative bacteria (P. aeruginosa and E. coli) was higher than that of Gram-positive bacteria (S. aureus). Finally, adding TiO2 NP to the films accelerated the healing process of the created wounds in a murine model, compared to control and CS-treated groups. CONCLUSION: Films of TiO2 NP and CS have suitable properties to be used in wound healing and can be further used in the future to load drugs.

Preparation and Characterization of Docetaxel-PLGA Nanoparticles Coated with Folic Acid-chitosan Conjugate for Cancer Treatment.

The conjugation of chitosan (CS) and folic acid (FA) was prepared and used to coat PLGA nanoparticles (NPs) that are loaded with Docetaxel (DTX) to target cancer cells that have lower pH and overexpression of folate receptors in comparison to normal cells. Three formulations had been prepared to reach the highest loading capacity (LC%) and encapsulation efficiency (EE%) and to study the effect of the amount of FA-CS on the drug release. The sizes, charges, homogeneity, surface morphology, LC% and EE% of the NPs were determined. The NPs were characterized using FTIR and XRD. In vitro release profiles of DTX from PLGA NPs, at pH 5.5 and 7.4 were determined. Finally, in vitro cytotoxicity assay on three cancer cell lines (RPMI 2650, Calu-3, and A549) was studied. The sizes of the three formulations ranged between 250.3±1.7 and 356.3±17.7. All prepared formulations showed acceptable monodispersity with highly positive charges. The EE% was above 85% and the LC% ranged between 6-35%. The in vitro release of DTX show an inverse relation to the amounts of FA-CS used and the pH of the dissolution medium. Coated PLGA NPs showed a significant difference in RPMI 2650, Calu-3, and A549 cell viability in comparison to free DTX. The NPs components were safe and non-toxic to human cells. In conclusion, coating PLGA NPs with FA-CS may be used as a good carrier for chemotherapeutic agents that selectively target carcinogenic tissues.

CSF Kappa Free Light Chains: Cutoff Validation for Diagnosing Multiple Sclerosis.

OBJECTIVE: To determine and validate a cerebrospinal fluid (CSF) κ (KCSF) value statistically comparable to detection of CSF-specific oligoclonal bands (OCB) to support the diagnosis of multiple sclerosis (MS). PATIENTS AND METHODS: A total of 702 retrospective and 657 prospective paired CSF/serum samples from residual waste samples of physician-ordered OCB tests were obtained and tested for KCSF at Mayo Clinic. Charts were reviewed by a neurologist blinded to KCSF results. Specificity and sensitivity for MS diagnosis were evaluated to establish a diagnostic cutoff value for KCSF in the retrospective cohort and then validated in the prospective cohort. RESULTS: Retrospective and prospective subgroups, respectively, included MS (n=85, 70), non-MS (n=615, 585), and undetermined diagnosis (excluded, n=2, 2). The retrospective data established a KCSF cutoff value of 0.1 mg/dL to be comparable to OCB testing. In the retrospective subgroup, KCSF vs OCB sensitivities for diagnosis of MS were 68.2% vs 75.0% (P=.08) and specificities were 86.1% vs 87.6% (P=.27). The KCSF area under the receiver operating characteristic curve was 0.772 (95% CI, 0.720 to 0.824), and for OCB was 0.813 (95% CI, 0.764 to 0.861). The prospective cohort was then used to validate the diagnostic KCSF value of 0.1 mg/dL; KCSF vs OCB sensitivities were 78.6% for both (P>.99) and specificities were 87.1% vs 89.4% (P=.09). CONCLUSION: The KCSF value of 0.1 mg/dL is a valid alternative to OCB testing, offering a standardized quantitative measure, eliminating human error, reducing cost and turnaround time, with no significant difference in sensitivity and specificity. This study provides class I evidence that a KCSF value of 0.1 mg/dL can be used in place of OCB testing to support the diagnosis of MS.

Polymeric Nanoparticles for Inhaled Vaccines.

Many recent studies focus on the pulmonary delivery of vaccines as it is needle-free, safe, and effective. Inhaled vaccines enhance systemic and mucosal immunization but still faces many limitations that can be resolved using polymeric nanoparticles (PNPs). This review focuses on the use of properties of PNPs, specifically chitosan and PLGA to be used in the delivery of vaccines by inhalation. It also aims to highlight that PNPs have adjuvant properties by themselves that induce cellular and humeral immunogenicity. Further, different factors influence the behavior of PNP in vivo such as size, morphology, and charge are discussed. Finally, some of the primary challenges facing PNPs are reviewed including formulation instability, reproducibility, device-related factors, patient-related factors, and industrial-level scale-up. Herein, the most important variables of PNPs that shall be defined in any PNPs to be used for pulmonary delivery are defined. Further, this study focuses on the most popular polymers used for this purpose.

The Pattern of Thyroid Malignancy and Its Associated Characteristics Among United Arab Emirates Population With More Focus on Patients in the Bethesda III Category.

INTRODUCTION: Thyroid cancer is the most prevalent endocrine cancer worldwide. It is the second most common type of cancer among United Arab Emirates (UAE) women and ranks as the sixth most common type of cancer overall among the UAE population. There are limited studies in the UAE related to thyroid malignancy. This study aimed to determine the pattern of thyroid malignancy among the UAE population and its associated characteristics, with more emphasis on patients categorized as Bethesda III by cytopathology, and furthermore, to determine the significance of advanced diagnostic methods in the assessment of thyroid nodules. METHODS: A retrospective review of the electronic medical charts of adult patients (age 18 and above) who were diagnosed with a thyroid nodule by ultrasound during the years 2019 and 2020. It is a comparative study of different variables associated with thyroid nodules and thyroid malignancy. RESULTS: A total of 1072 patients were diagnosed with thyroid nodules upon initial ultrasound. We had 174 patients diagnosed with thyroid malignancy, constituting 16% (95% CI 0.14-0.19) of the total study population. 78% of the thyroid malignancy patients were women as compared to men, and this difference was statistically significant (p=0.042). Non-UAE nationals comprised 61% of the population diagnosed with thyroid malignancy (95% CI 1.37-2.68). Malignancy was found to be more common in patients with multinodular goiter, in the 30 to 39-year age group, and in patients with high ultrasound and Bethesda grades. From the total study population, 140 patients had cytology reports in the Bethesda III category. Thyroid malignancy was found in 30 patients with Bethesda III, and this comprised 17% of the total population who were diagnosed with thyroid malignancy. CONCLUSION: Despite being a single-center study, it highlights the percentage of thyroid malignancy and its associated factors among the UAE population. Thyroid ultrasound grading and Bethesda classification guide physicians in risk stratification, but it remains challenging in patients who fall into the Bethesda III category. Intervention versus regular follow-up should not depend on a single value but on the overall clinical picture and the use of advanced diagnostic methods.

The scavenger receptor CD36 contributes to the neurotoxicity of bone marrow-derived monocytes through peroxynitrite production.

CD36, a class B scavenger receptor present in microglia, endothelium and leukocytes, plays a key role in ischemic brain injury by promoting the expression of inflammatory genes and production of reactive oxygen species (ROS). However, it is not known whether ischemic brain damage is mediated by CD36 activation in resident brain cells, i.e., microglia, or by blood-borne cells that infiltrate the brain. To address this question, we studied oxygen-glucose deprivation (OGD) in hippocampal slice cultures, a model of ischemic injury that does not involve cells extrinsic to the brain. We found that CD36 gene knockout does not afford protection of hippocampal slices to OGD-induced cytotoxicity. In contrast, immunoactivated bone marrow-derived monocytes-macrophages (BMM) from wild type (WT) mice trigger hippocampal damage when incubated with brain slices via a mechanism that is prevented in CD36-/- BMM. The neurotoxic activity of CD36+/+ BMM was attributed to reactive oxygen species (ROS) since it was concomitant with increased ROS production and could be prevented by treatment with a selective ROS scavenger, MnTBAP, or a peroxynitrite decomposition catalyst, FeTPPS. Importantly, ROS production and accumulation 3-nitrotyrosine in hippocampal proteins (a hallmark of peroxynitrite production) was significantly dampened in immunoactivated CD36-/- BMM, whereas production of NO-derived metabolites (nitrite and nitrate) was unaltered. We conclude that CD36 signaling may not contribute to injury induced by OGD in the brain itself but is involved in the neurotoxicity mediated by activated BMM. These findings are consistent with the hypothesis that CD36 in infiltrating inflammatory cells drives peroxynitrite-mediated ischemic brain damage. Accordingly, targeting CD36 in the vascular compartment may protect against neurotoxicity in the ischemic brain.

Inducible nitric oxide synthase provides protection against injury-induced thrombosis in female mice.

Nitric oxide (NO) is an important vasoactive molecule produced by three NO synthase (NOS) enzymes: neuronal (nNOS), inducible (iNOS), and endothelial NOS (eNOS). While eNOS contributes to blood vessel dilation that protects against the development of hypertension, iNOS has been primarily implicated as a disease-promoting isoform during atherogenesis. Despite this, iNOS may play a physiological role via the modulation of cyclooxygenase and thromboregulatory eicosanoid production. Herein, we examined the role of iNOS in a murine model of thrombosis. Blood flow was measured in carotid arteries of male and female wild-type (WT) and iNOS-deficient mice following ferric chloride-induced thrombosis. Female WT mice were more resistant to thrombotic occlusion than male counterparts but became more susceptible upon iNOS deletion. In contrast, male mice (with and without iNOS deletion) were equally susceptible to thrombosis. Deletion of iNOS was not associated with a change in the balance of thromboxane A(2) (TxA(2)) or antithrombotic prostacyclin (PGI(2)). Compared with male counterparts, female WT mice exhibited increased urinary nitrite and nitrate levels and enhanced ex vivo induction of iNOS in hearts and aortas. Our findings suggest that iNOS-derived NO in female WT mice may attenuate the effects of vascular injury. Thus, although iNOS is detrimental during atherogenesis, physiological iNOS levels may contribute to providing protection against thrombotic occlusion, a phenomenon that may be enhanced in female mice.

Strabismus Surgery Outcomes and Their Determinants in Patients with Chronic Sixth Nerve Palsy.

PURPOSE: Patients with chronic sixth nerve palsy (CSNP) comprise a heterogeneous population, and the optimal surgical solution remains uncertain. Here, we present the success rate and factors associated with the success of strabismus surgeries for CSNP. METHODS: This was a retrospective cohort study of patients with strabismus due to CSNP operated on between 2015 and 2019 in a tertiary eye hospital in central Saudi Arabia. Surgical success was defined as a horizontal deviation of ≤10 prism diopters (PDs) assessed at least 12 months after surgery. Differences between groups with respect to the primary outcome were assessed. RESULTS: Fifty-five patients were analyzed with a median follow-up of 24 (range 12-48) months. Superior rectus and inferior rectus transposition (34.5%) and medial rectus recession with lateral rectus resection (32.7%) were the main surgeries performed. The overall success rate was 67.3% (95% confidence interval 54.9-79.7). Bilateral CSNP (P = 0.05), a higher preoperative angle of deviation (P = 0.002), or a greater degree of preoperative limitation of abduction (P = 0.012), but not the type of surgery (P = 0.09), were more likely to result in an under-corrected outcome of >10 PD. The preoperative deviation angle showed a trend toward being associated with a poor outcome after surgery (P = 0.06). Six patients with high-angle deviation before surgery required second surgery. CONCLUSION: While the surgical procedure does not impact outcomes, the severity of preoperative horizontal deviation might impact surgical success and the need for reoperation. Patients with severe CSNP should be counseled appropriately about the chances of surgical success and the potential need for further interventions.

Physical evidence for substrate binding in preventing cyclooxygenase inactivation under nitrative stress.

Prostaglandin biosynthesis is catalyzed by two spatially and functionally distinct active sites in cyclooxygenase (COX) enzymes. Despite the crucial role of COXs in biology, molecular details regarding the function and regulation of these enzymes are incompletely defined. Reactive nitrogen species, formed during oxidative stress, produce modifications that alter COX functionalities and prostaglandin biosynthesis. We previously established that COX-1 undergoes selective nitration on Tyr385 via a mechanism that requires the presence of bound heme cofactor. As this is a critical residue for COX-1 catalysis, nitration at this site results in enzyme inactivation. We now show that occupancy of the COX-1 active site with substrate protects against Tyr385 nitration and redirects nitration to alternative Tyr residues on COX-1, preserving catalytic activity. This study reveals a novel role for the substrate in protecting COX-1 from inactivation by nitration in pathophysiological settings.

Inducible nitric oxide synthase gene deletion exaggerates MAPK-mediated cyclooxygenase-2 induction by inflammatory stimuli.

Cyclooxygenase (COX)-2 and inducible nitric oxide (NO) synthase (iNOS) are responsive to a wide array of inflammatory stimuli, have been localized to vascular smooth muscle cells (SMCs), and are intimately linked to the progression of vascular disease, including atherosclerotic lesion formation. We and others have shown that the production and subsequent impact of COX products appear to be correlative with the status of NO synthesis. This study examined the impact of inflammation-driven NO production on COX-2 expression in SMCs. Concurrent stimulation of quiescent rat aortic SMCs with lipopolysaccharide (LPS) and interferon (IFN)-gamma increased COX-2, iNOS, and nitrite production. Pharmacological inhibition of NO synthase (N(G)-monomethyl-l-arginine) concentration- and time-dependently magnified LPS + IFN-gamma-mediated COX-2 mRNA and protein induction in a cGMP-independent manner. COX-2 induction was associated with activation of the ERK, p38, and JNK mitogen-activated protein kinase (MAPK) pathways. Interestingly, NO synthase inhibition enhanced ERK, p38, and to a lesser extent JNK phosphorylation but suppressed MAPK phosphatase (MKP)-1 induction in response to LPS + IFN-gamma. Similarly, the exposure of SMCs from iNOS(-/-) mice to LPS + IFN-gamma produced an enhancement of COX-2 induction, p38, and JNK phosphorylation and an attenuated upregulation of MKP-1 versus their wild-type counterparts. Taken together, our data indicate that NO, in part derived from iNOS, negatively regulates the immediate early induction of COX-2 in response to inflammatory stimuli.