Objective: Mitochondrial dysfunction is closely linked to chronic disorders. This study aims to explore the correlation between pediatric anemia and mitochondrial markers, specifically fibroblast growth factor 21 (FGF21), growth/differentiation factor 15 (GDF-15), and nitric oxide synthase (eNOS). Method: This study included 66 children, with 34 diagnosed with anemia and 32 in the healthy control group. Statistically significant biomarkers were determined through cutoff levels. Results: Among the participants, 34 children were classified as anemic, while 32 were categorized as healthy. The study revealed that FGF21 levels ≥ 0.745 pg/mL and eNOS levels ≥ 1.265 µg/mL predicted anemia. Hemoglobin levels exhibited a negative correlation with FGF21 (r = -0.381; p = 0.002) and eNOS levels (r = -0.462; p < 0.001). Furthermore, a significant negative correlation was observed between GDF-15 and ferritin (r = -0.311; p = 0.019), while eNOS levels correlated positively with folate (r = 0.313; p = 0.019). Conclusions: Anemia induced elevated mitochondrial biomarkers; FGF21 and eNOS levels. The findings suggest that the long-term ramifications of anemia in childhood may be associated with mitochondrial dysfunction.

In response to mechanical loading of bone, osteocytes produce nitric oxide (NO•) and decrease sclerostin protein expression, leading to an increase in bone mass. However, it is unclear whether NO• production and sclerostin protein loss are mechanistically linked, and, if so, the nature of their hierarchical relationship within an established mechano-transduction pathway. Prior work showed that following fluid-shear stress (FSS), osteocytes produce NOX2-derived reactive oxygen species, inducing calcium (Ca2+) influx. Increased intracellular Ca2+ results in calcium-calmodulin dependent protein kinase II (CaMKII) activation, which regulates the lysosomal degradation of sclerostin protein. Here, we extend our discoveries, identifying NO• as a regulator of sclerostin degradation downstream of mechano-activated CaMKII. Pharmacological inhibition of nitric oxide synthase (NOS) activity in Ocy454 osteocyte-like cells prevented FSS-induced sclerostin protein loss. Conversely, short-term treatment with a NO• donor in Ocy454 cells or isolated murine long bones was sufficient to induce the rapid decrease in sclerostin protein abundance, independent of changes in Sost gene expression. Ocy454 cells express all three NOS genes, and transfection with siRNAs targeting eNOS/Nos3 was sufficient to prevent FSS-induced loss of sclerostin protein, while siRNAs targeting iNOS/Nos2 mildly blunted the loss of sclerostin but did not reach statistical significance. Similarly, siRNAs targeting both eNOS/Nos3 and iNOS/Nos2 prevented FSS-induced NO• production. Together, these data show iNOS/Nos2 and eNOS/Nos3 are the primary producers of FSS-dependent NO•, and that NO• is necessary and sufficient for sclerostin protein control. Further, selective inhibition of elements within this sclerostin-controlling mechano-transduction pathway indicated that NO• production occurs downstream of CaMKII activation. Targeting Camk2d and Camk2g with siRNA in Ocy454 cells prevented NO• production following FSS, indicating that CaMKII is needed for NO• production. However, NO• donation (1min) resulted in a significant increase in CaMKII activation, suggesting that NO• may have the ability to tune CaMKII response. Together, these data support that CaMKII is necessary for, and may be modulated by NO•, and that the interaction of these two signals is involved in the control of sclerostin protein abundance, consistent with a role in bone anabolic responses.

Glioblastoma (GBM) is the most common malignant primary brain tumor, resulting in poor survival despite aggressive therapies. GBM is characterized by a highly heterogeneous and immunosuppressive tumor microenvironment (TME) made up predominantly of infiltrating peripheral immune cells. One significant immune cell type that contributes to glioma immune evasion is a population of immunosuppressive cells, termed myeloid-derived suppressor cells (MDSCs). Previous studies suggest that a subset of myeloid cells, expressing monocytic (M)-MDSC markers and dual expression of chemokine receptors CCR2 and CX3CR1, utilize CCR2 to infiltrate the TME. This study evaluated the mechanism of CCR2+/CX3CR1+ M-MDSC differentiation and T cell suppressive function in murine glioma models. We determined that bone marrow-derived CCR2+/CX3CR1+ cells adopt an immune suppressive cell phenotype when cultured with glioma-derived factors. Glioma secreted CSF1R ligands M-CSF and IL-34 were identified as key drivers of M-MDSC differentiation while adenosine and iNOS pathways were implicated in M-MDSC suppression of T cells. Mining a human GBM spatial RNAseq database revealed a variety of different pathways that M-MDSCs utilize to exert their suppressive function that are driven by complex niches within the microenvironment. These data provide a more comprehensive understanding of the mechanism of M-MDSCs in glioblastoma.

Cardiovascular functions in diabetes greatly depend on constitutive NOS (cNOS) activity. A comparative study of the effects of a steroid hormone ecdysterone and enalapril, an ACE inhibitor widely used to treat cardiac disorders on cNOS, inducible NOS (iNOS), xanthine oxidoreductase (XOR) activity, RNS, ROS, and lipid peroxidation in heart tissue in experimental diabetes was conducted. The rat model of diabetes was established by streptozotocin injection. NOS activity, NO2-, NO3-, uric acid, nitrosothiols, hydroperoxide, superoxide, and diene conjugate formation were studied spectrophotomerically. In diabetes, cNOS downregulation correlated with a dramatic fall of NO2- production and ~4.5-fold elevation of nitrosothiols, which agreed with a steep rise of iNOS activity, while NO3- remained close to control. Dramatic activation of XOR was observed, which correlated with the elevation of both superoxide production and nitrate reductase activity and resulted in strong lipid peroxidation. Ecdysterone and enalapril differently affected RNS metabolism. Ecdysterone moderately restored cNOS but strongly suppressed iNOS, which resulted in the reduction of NO3-, but full restoration of NO2- production. Enalapril better restored cNOS but less effectively suppressed iNOS, which promoted NO3- formation. Both drugs similarly inhibited XOR, which equally alleviated oxidative stress and lipid peroxidation. The synergistic action of iNOS and XOR was a plausible explanation for strong lipid peroxidation, abolished by the inhibition of iNOS and XOR by ecdysterone or enalapril. Complementary effects of ecdysterone and enalapril on cNOS, iNOS, and RNS are a promising basis for their combined use in the treatment of cardiovascular disorders caused by cNOS dysfunction in diabetes.

UNASSIGNED: The objective of this study was to describe predictors of loss of ambulation in Duchenne muscular dystrophy (DMD).
UNASSIGNED: This systematic review and meta-analysis included searches of MEDLINE ALL, Embase, and the Cochrane Database of Systematic Reviews from January 1, 2000, to December 31, 2022, for predictors of loss of ambulation in DMD. Search terms included \"Duchenne muscular dystrophy\" as a Medical Subject Heading or free text term, in combination with variations of the term \"predictor\". Risk of bias was assessed using the Newcastle-Ottawa Scale. We performed meta-analysis pooling of hazard ratios of the effects of glucocorticoids (vs. no glucocorticoid therapy) by fitting a common-effect inverse-variance model.
UNASSIGNED: The bibliographic searches resulted in the inclusion of 45 studies of children and adults with DMD from 17 countries across Europe, Asia, and North America. Glucocorticoid therapy was associated with delayed loss of ambulation (overall meta-analysis HR deflazacort/prednisone/prednisolone: 0.44 [95% CI: 0.40-0.48]) (n = 25 studies). Earlier onset of first signs or symptoms, earlier loss of developmental milestones, lower baseline 6MWT (i.e.,<350 vs. ≥350 metres and <330 vs. ≥330 metres), and lower baseline NSAA were associated with earlier loss of ambulation (n = 5 studies). Deletion of exons 3-7, proximal mutations (upstream intron 44), single exon 45 deletions, and mutations amenable of skipping exon 8, exon 44, and exon 53, were associated with prolonged ambulation; distal mutations (intron 44 and downstream), deletion of exons 49-50, and mutations amenable of skipping exon 45, and exon 51 were associated with earlier loss of ambulation (n = 13 studies). Specific single-nucleotide polymorphisms in CD40 gene rs1883832, LTBP4 gene rs10880, SPP1 gene rs2835709 and rs11730582, and TCTEX1D1 gene rs1060575 (n = 7 studies), as well as race/ethnicity and level of family/patient deprivation (n = 3 studies), were associated with loss of ambulation. Treatment with ataluren (n = 2 studies) and eteplirsen (n = 3 studies) were associated with prolonged ambulation. Magnetic resonance biomarkers (MRI and MRS) were identified as significant predictors of loss of ambulation (n = 6 studies). In total, 33% of studies exhibited some risk of bias.
UNASSIGNED: Our synthesis of predictors of loss of ambulation in DMD contributes to the understanding the natural history of disease and informs the design of new trials of novel therapies targeting this heavily burdened patient population.

OBJECTIVE: The aim of the study was to evaluate the hemodynamic and RA system effects of the oral administration of the clinical dose of beraprost for feline CKD in healthy cats, and also to examine whether NOS inhibition reversed them.
METHODS: A placebo-controlled pharmacological sequential design study was carried out to assess the plasma aldosterone and renin concentrations (PAC and PRC), blood pressure, heart rate, and exploratorily to estimate renal plasma flow (RPF) and renal vascular resistance (RVR) with simplified methods.
RESULTS: Beraprost reduced PAC when compared to the placebo (p < 0.05); this was reversed when NOS inhibitor NG-nitro-L-arginine methyl ester (L-NAME) was added to the beraprost treatment (p < 0.01). No differences in the PRC or hemodynamic parameters were detected between beraprost and the placebo. The correlation ratios (η2) showed opposite relationships between beraprost and the added L-NAME effects on PAC, mean blood pressure (MBP), heart rate, estimated RPF (p < 0.001), estimated RVR (p < 0.01), and PRC (p < 0.05).
CONCLUSIONS: In healthy cats, the clinical dose of beraprost suppresses PAC, which can be reversed by the inhibition of NOS.

The functional importance of nitric oxide (NO) in the fields of immunology concerning its antimicrobial, anti-tumoral, anti-inflammatory, and immunosuppressive effects have made it inevitable to study its secretion from various cells. Nitrogen oxide synthase (NOS) is the enzyme responsible for synthesizing NO and its three isoforms function in a cell-dependent manner. NO is oxidized rapidly to Reactive nitrogen oxide species (RNOS) through which the roles of NO are being carried out. One of the major immune cells secreting NO is myeloid-derived suppressor cells (MDSCs). The function of these MDSCs in the suppression of T-cell proliferation as well as T-cell differentiation is found to be dependent on NO secretion. Apart from T-cell suppressive activity, NO is also known to interfere with natural killer (NK) cell functions. A convenient method to estimate NO secretion is by using Griess reagent named after Johann Peter Griess. In this method, NO reacts with the reagents to form a colored azo dye detectable using a microplate reader at a wavelength of 548nm. In this chapter, we summarized the detailed method of estimating NO from MDSCs by the Griess method.

Sodium valproate (VPA), a histone deacetylase (HDAC) inhibitor, could be a promising candidate to treat acute myocardial infarction (AMI). In this study, AMI was induced in New Zealand White rabbits by occluding the left circumflex coronary artery for 1 h, followed by reperfusion. The animals were distributed into three experimental groups: the sham-operated group (SHAM), the AMI group and the AMI + VPA group (AMI treated with VPA 500 mg/kg/day). After 5 weeks, abdominal aorta was removed and used for isometric recording of tension in organ baths or protein expression by Western blot, and plasma for the determination of nitrate/nitrite (NOx) levels by colorimetric assay. Our results indicated that AMI induced a reduction of the endothelium-dependent response to acetylcholine without modifying the endothelium-independent response to sodium nitroprusside, leading to endothelial dysfunction. VPA treatment reversed AMI-induced endothelial dysfunction and even increased NO sensitivity in vascular smooth muscle. This response was consistent with an antioxidant effect of VPA, as it was able to reverse the superoxide dismutase 1 (SOD 1) down-regulation induced by AMI. Our experiments also ruled out that the VPA mechanism was related to eNOS, iNOS, sGC and arginase expression or changes in NOx plasma levels. Therefore, we conclude that VPA improves vasodilation by increasing NO bioavailability, likely due to its antioxidant effect. Since endothelial dysfunction was closely related to AMI, VPA treatment could increase aortic blood flow, making it a potential agent in reperfusion therapy that can prevent the vascular damage.

It is obvious that the oxidation process is an undeniable fact and when it comes to aging, one of the first solutions that come to mind is natural products. When it comes to natural products, both plants and bee products play an important, almost combative role against oxidation. For this purpose, natural products of both plant and animal origin were considered together in our study: Linden, green tea, aronia, wild grapes, myrtle, blueberries and basil, honey, pollen and propolis. Total phenolic content values of the extracts ranged between 49.28 and 3859.06 mg gallic acid equivalent/100 g, and propolis, green tea, chestnut flower and aronia samples were found to have the highest values. When looking at the NOS inhibition potential, it was determined that propolis, pollen and aronia samples had the highest percentage inhibition values of 98.11, 92.29, 83.44, respectively. Antioxidant activities of methanolic extracts were investigated using iron(III) reducing/antioxidant capacity (FRAP), 2,2-Diphenyl-1-picrylhydrazyl (DPPH) radical scavenging activity test and NOS inhibition tests. The phenolic composition of methanolic extracts was tested using the RP-HPLC-UV (high-performance liquid chromatographic method with ultraviolet) method with 19 phenolic standards.

OBJECTIVE: The objective of this study is to explore the impact and underlying mechanism of Scutellaria baicalensis Georgi stem and leaf flavonoids (SSFs) on cognitive impairment caused by intracerebroventricular injection of okadaic acid (OA) in rats.
METHODS: An experimental model of Alzheimer\'s disease (AD) was induced in rats by intracerebroventricular injection of OA, resulting in memory impairment. The Morris water maze test was employed to confirm the successful establishment of the memory impairment model. The rats that exhibited significant memory impairment were randomly divided into different groups, including a model group, three SSFs dose groups (25, 50, and 100 mg/kg), and a positive control group treated with Ginkgo biloba tablets (GLT) at a dose of 200 mg/kg. To evaluate the learning and memory abilities of the rats, the Morris water maze test was conducted. Hematoxylin-eosin (HE) staining was used to observe any morphological changes in neurons. Immunohistochemistry (IHC) was performed to measure the expression of choline acetyltransferase (ChAT) protein. Western blotting (WB) was utilized to assess the phosphorylation levels of tau protein at Ser262 and Ser396. The activities of inducible nitric oxide synthase (iNOS) and constitutive nitric oxide synthase (cNOS) were quantified using ultraviolet spectrophotometry. The levels of inflammatory factors, including interleukin-1β (IL-1β), tumor necrosis factor-α (TNF-α), and interleukin-6 (IL-6), were measured using ELISA.
RESULTS: In rats, the administration of OA via intracerebroventricular injection resulted in cognitive impairment, neuropathological changes, and alterations in protein expression and activity levels. Specifically, the protein expression of ChAT was significantly reduced (P<0.01), while the phosphorylation levels of tau protein at Ser262 and Ser396 were significantly increased (P<0.01). Moreover, iNOS activity in the hippocampus and cerebral cortex exhibited a significant increase (P<0.01), whereas cNOS activity showed a decrease (P<0.05). Furthermore, the levels of IL-1β and TNF-α in the cerebral cortex were elevated (P<0.01), while the level of IL-6 was decreased (P<0.05). The administration of three doses of SSFs and GLT to rats exhibited varying degrees of improvement in the aforementioned pathological alterations induced by OA.
CONCLUSIONS: SSFs demonstrated the ability to enhance cognitive function and mitigate memory deficits in rats following intracerebroventricular injection of OA. This beneficial effect may be attributed to the modulation of ChAT protein expression, tau hyperphosphorylation, NOS activity, and inflammatory cytokine levels by SSFs.