Active Vitamin D3 Research Articles

A pharmacovigilance study on clinical factors of active vitamin D3 analog-related acute kidney injury using the Japanese Adverse Drug Event Report Database

Acute kidney injury (AKI) due to vitamin D therapy for osteoporosis is encountered in clinical practice, but epidemiological studies are scarce. We aimed to determine the association between AKI and vitamin D therapy and to identify risk factors for AKI using the Japanese Adverse Drug Event Report database. We used reporting odds ratios (RORs) to detect signals and evaluate risk factors using multiple logistic regression analysis. Among 298,891 reports from April 2004 to September 2023, 1071 implicated active vitamin D3 analogs as suspect drugs for adverse events. There was a significant association between AKI and active vitamin D3 analogs (ROR [95% confidence interval {CI}], eldecalcitol: 16.75 [14.23–19.72], P < 0.001; alfacalcidol: 5.29 [4.07–6.87], P < 0.001; calcitriol: 4.46 [1.88–10.59], P < 0.001). The median duration of administration before AKI onset was 15.4 weeks. Multiple logistic regression analysis showed a significant association between AKI and age ≥ 70 years (odds ratio [95% CI], 1.47 [1.04–2.07]; P = 0.028), weight < 50 kg (1.55 [1.12–2.13]; P = 0.007), hypertension (1.90 [1.42–2.54]; P < 0.001), and concomitant use of nonsteroidal anti-inflammatory drugs (1.58 [1.10–2.25], P = 0.012) and magnesium oxide (1.96 [1.38–2.78]; P < 0.001). Our results suggest that active vitamin D3 analogs are associated with AKI development. Physicians prescribing these medications to patients with risk factors should consider the possibility of AKI, especially during the first 6 months.

The epigenetic effects of glucocorticoids, sex hormones and vitamin D as steroidal hormones in rheumatic musculoskeletal diseases.

Chronic rheumatological diseases are multifactorial conditions in which both the neuroendocrine hormone pathway, including cortisol, sex hormones and active vitamin D3 (calcitriol), all deriving from cholesterol, and the epigenetic modifications that they cause play an important role. In fact, epigenetics modulates the function of the DNA of immune cells, through three main mechanisms: DNA methylation, modifications to the histones that make up chromatin and production of non-coding RNAs (microRNA - miRNA). In this narrative review, the main data regarding the epigenetic modifications induced by cortisol, 17β-oestradiol, progesterone, testosterone and calcitriol on immune cells were collected, discussing how these can interfere in the predisposition and course of chronic rheumatological diseases (i.e. rheumatoid arthritis, systemic lupus erythematosus, systemic sclerosis). An ever-increasing number of miRNAs have been identified, which are produced by neuroendocrine hormones and can influence the inflammatory-fibrotic response at various levels. Concerning the involvements of the neuro-endocrine-immunology within the pathophysiology of rheumatic diseases, the epigenetic effects induced by steroid hormones must be taken into consideration to evaluate their impact on the progression of the single condition and even inside the single patient.

Vitamin D, the Sunshine Molecule That Makes Us Strong: What Does Its Current Global Deficiency Imply?

Vitamin D3 deficiency and insufficiency are becoming a common global issue for us, especially in the most industrially developed countries. The only acknowledged activity of vitamin D3 in vertebrates is to promote the absorption of calcium and, therefore, allow for the mineralization of bones. Accordingly, its deficiency is associated with diseases such as rickets. Other numerous vital functions associated with vitamin D3 are yet to be considered, and the function of vitamin D2 in plants is unknown. Thus, 100 years after its discovery, the importance of vitamin D still seems to be unacknowledged (except for rickets), with little attention given to its decrease throughout the world. In this review, I suggest that vitamin D deficiency and insufficiency may be linked to the westernized lifestyle in more developed countries. Furthermore, I suggest that, rather than the calcemic activity, the main function of vitamin D is, in general, that of strengthening living organisms. I conclude with the hypothesis that vitamin D deficiency may represent a marker for a greater risk of chronic inflammatory diseases and a shorter life expectancy.

Palopegteriparatide Treatment Improves Renal Function in Adults with Chronic Hypoparathyroidism: 1-Year Results from the Phase3 PaTHway Trial.

Individuals with chronic hypoparathyroidism managed with conventional therapy (active vitaminD and calcium) have an increased risk for renal dysfunction versus age- and sex-matched controls. Treatments that replace the physiologic effects of parathyroid hormone (PTH) while reducing the need for conventional therapy may help prevent a decline in renal function in this population. This post hoc analysis examined the impact of palopegteriparatide treatment on renal function in adults with chronic hypoparathyroidism. PaTHway is a phase3 trial of palopegteriparatide in adults with chronic hypoparathyroidism that included a randomized, double-blind, placebo-controlled 26-week period followed by an ongoing 156-week open-label extension (OLE) period. Changes in renal function over 52weeks (26weeks blinded + 26weeks OLE) were assessed using estimated glomerular filtration rate (eGFR). A subgroup analysis was performed with participants stratified by baseline eGFR < 60 or ≥ 60mL/min/1.73m2. At week52, over 95% (78/82) of participants remained enrolled in the OLE and of those, 86% maintained normocalcemia and 95% achieved independence from conventional therapy (no active vitaminD and ≤ 600mg/day of calcium), with none requiring active vitaminD. Treatment with palopegteriparatide over 52weeks resulted in a mean (SD) increase in eGFR of 9.3 (11.7)mL/min/1.73m2 from baseline (P < 0.0001) and 43% of participants had an increase ≥ 10mL/min/1.73m2. In participants with baseline eGFR < 60mL/min/1.73m2, 52weeks of treatment with palopegteriparatide resulted in a mean (SD) increase of 11.5 (11.3) mL/min/1.73m2 (P < 0.001). One case of nephrolithiasis was reported for a participant in the placebo group during blinded treatment; none were reported through week52 with palopegteriparatide. In this post hoc analysis of the PaTHway trial, palopegteriparatide treatment was associated with significantly improved eGFR at week52 in addition to previously reported maintenance and normalization of serum and urine biochemistries. Further investigation of palopegteriparatide for the preservation of renal function in hypoparathyroidism is warranted. ClinicalTrials.gov NCT04701203.

Electro-scalp acupuncture regulates the expression of CYP27a1/b1, CYP24a and related inflammatory cytokines in ischemic cortex of rats with middle cerebral artery occlusion.

To observe the effect of electro-scalp acupuncture (ESA) on the expression of cytochrome P450a1/b1 (CYP27a1/b1), cytochrome P45024a (CYP24a), signal transducer and activator of transcription (STAT)4, STAT6, tumor necrosis factor-α (TNF-α), interleukin (IL)-1β and IL-4 in ischemic cerebral cortex of rats with acute ischemic stroke, so as to explore its mechanism in alleviating inflammatory reaction of ischemic stroke. Sixty SD rats were randomly divided into sham-operation, model, vitamin D3 and ESA groups, with 15 rats in each group. The middle cerebral artery occlusion rat model was established with thread ligation according to Zea-Longa's method. Rats in the vitamin D3 group were given 1, 25-VitD3 solution (3 ng·100 g-1·d-1) by gavage, once daily for 7 days. Rats in the ESA group were treated at bilateral anterior parietotemporal slash (MS6) with ESA (2 Hz/100 Hz, 1 mA), 30 min a day for 7 days. Before and after interventions, the neurological deficit score and neurobehavioral score were evaluated. TTC staining was used to detect the volume of cerebral infarction in rats. The positive expressions of CYP24a, CYP27a1 and CYP27b1 in the cerebral cortex of ischemic area were detected by immunofluorescence. The mRNA expressions of STAT4 and STAT6 in the cerebral cortex of ischemic area were detected by quantitative real-time PCR. The protein expression levels of TNF-α, IL-1β and IL-4 in the cerebral cortex of ischemic area were detected by Western blot. Compared with the sham-operation group, the neurological deficit score, neurobehavioral score, the percentage of cerebral infarction volume, the positive expression level of CYP24a and mRNA expression level of STAT4, protein expression levels of TNF-α and IL-1β in cerebral cortex were increased (P<0.01), while the positive expression levels of CYP27a1/b1 and STAT6 mRNA, protein expression level of IL-4 were decreased (P<0.01) in the model group. After the treatment and compared with the model group, the neurological deficit score, neurobehavioral score, the percentage of cerebral infarction volume, the positive expression level of CYP24a and mRNA expression level of STAT4, protein expression levels of TNF-α and IL-1β in cerebral cortex were decreased (P<0.01), while the positive expression levels of CYP27a1/b1 and STAT6 mRNA expression level, protein expression level of IL-4 were increased (P<0.01) in the ESA and vitamin D3 groups. ESA can alleviate the inflammatory response in ischemic stroke, which maybe related to its function in regulating the balance between CYP27a1/b1 and CYP24a, converting vitamin D into active vitamin D3, inhibiting vitamin D3 degradation, and regulating Th1/Th2 balance.

The Antimicrobial Activity of Combination of Vitamin D3 and Omega-3 against Pathogenic Microorganisms.

Food supplements such as vitamin D3 and omega-3 have a significant role in activating the immune system and impacting the diversity of gut microbiota; thus, controlling the growth of invading pathogens indirectly. This study aims to evaluate the direct antimicrobial activity of vitamin D3 and omega- 3 individually, combined together, and combined with low concentrations of gentamicin or amphotericin B against selected pathogenic microorganisms. In addition, this study hypothesizes the potential antimicrobial mechanism and recommends suitable studies to be conducted. Minimum inhibitory concentration of different serial dilutions of vitamin D3 [0.7μg/mL-83.3μg/mL] or omega-3 [0.7mg/mL-100mg/mL] or combined [vitamin D3:1.3μg/mL-83.3μg/mL and omega-3:1.56mg/mL-100mg/mL] with/without antibiotic have been investigated on Escherichia coli, Staphylococcus aureus, Pseudomonas aeruginosa and Candida albicans using check board technique. The highest concentration of vitamin D3 [83.3 μg/mL] demonstrated a complete eradication of the tested microorganisms. Conversely, omega-3 had a lower effect on them. The highest concentration of combining vitamin D3 and omega-3 with/without gentamicin resulted in a complete eradication of the S. aureus, E. coli and P. aeruginosa with a 6.8 to 7 log reduction. On the other hand, C. albicans was inhibited when using vitamin D3 [83.3 μg/mL] or when this concentration is combined with 100mg/mL of omega-3. However, when these two concentrations were added to amphotericin B the log reduction dropped to 0.45 suggesting antagonistic effect. These findings suggested that, unlike omega 3, vitamin D3 possesses good antimicrobial effects against pathogenic microorganisms. The combination of the studied food supplement showed enhanced microbial inhibition at high concentration, while they had antagonistic effect when combined with amphotericin B and applied on C. albicans combined. Further studies on the exact antimicrobial mechanism are still required to understand the measured data here.

Protective Effects of Dietary Vitamin D3, Turmeric Powder, and Their Combination against Gasoline Intoxication in Rats.

The inhalation of gasoline vapors (GV) is associated with developing various pathologies. Particularly, oil refinery and gas station workers are at a greater risk of developing lung cancer, kidney cancer, bladder cancer, and hematological disorders, including acute myeloid leukemia. Therefore, preventing the harmful effects of GV and alleviating their consequences appear to be important and timely issues. In this study, we investigated the potential of vitamin D3, turmeric powder, and their combination to ameliorate the toxicity of gasoline fumes in rats. Separate groups of animals fed with a standard rodent diet, with or without the supplementation of vitamin D3 (750 IU/kg body weight) and/or turmeric powder (0.5%, w/w, in food), were untreated or treated with GV (11.5 ± 1.3 cm3/h/m3/day) for 30, 60, or 90 days. Changes in the body weight were monitored weekly. Histological, biochemical, and hematological parameters were determined at the end of each treatment period. While the exposure of rats to GV resulted in a time-dependent reduction in body weight, supplementation with vitamin D3, but not with turmeric root powder or their combination, partially prevented weight loss. Macroscopical and histological analyses showed pronounced time-dependent changes in the organs and tissues of GV-treated rats. These included alveolar wall collapse in the lungs, the destruction of the lobular structure and hepatocytolysis in the liver, the shrinkage and fragmentation of glomeruli in the kidneys, and the disorganization of the lymphoid follicles in the spleen. However, co-treatment with the nutritional supplements tested, especially vitamin D3, noticeably alleviated the above conditions. This was accompanied by a significant improvement in the blood chemistry and hematological parameters. Collectively, our results demonstrate that the harmful effects of environmental exposure to GV can be reduced upon supplementation of vitamin D3. The fact that the protective activity of vitamin D3 alone was higher than that of turmeric root powder or the combined treatment suggests that combinations of these supplements may not always be more beneficial than each agent applied separately.

Unveiling the Novel Benefits of Co-Administering Butyrate and Active Vitamin D3 in Mice Subjected to Chemotherapy-Induced Gut-Derived Pseudomonas aeruginosa Sepsis.

Cancer patients face increased susceptibility to invasive infections, primarily due to ulcerative lesions on mucosal surfaces and immune suppression resulting from chemotherapy. Pseudomonas aeruginosa (P. aeruginosa) bacteremia is notorious for its rapid progression into fatal sepsis, posing a significant threat to cancer patients, particularly those experiencing chemotherapy-induced neutropenia. This bacterial infection contributes significantly to morbidity and mortality rates among such individuals. Our latest report showed the mutually beneficial effects of postbiotic butyrate on 1,25-dihydroxyvitamin D3 (1,25D3)-controlled innate immunity during Salmonella colitis. Hence, we investigated the impact of butyrate and 1,25D3 on chemotherapy-induced gut-derived P. aeruginosa sepsis in mice. The chemotherapy-induced gut-derived P. aeruginosa sepsis model was established through oral administration of 1 × 107 CFU of the P. aeruginosa wild-type strain PAO1 in C57BL/6 mice undergoing chemotherapy. Throughout the infection process, mice were orally administered butyrate and/or 1,25D3. Our observations revealed that the combined action of butyrate and 1,25D3 led to a reduction in the severity of colitis and the invasion of P. aeruginosa into the liver and spleen of the mice. This reduction was attributed to an enhancement in the expression of defensive cytokines and antimicrobial peptides within the cecum, coupled with decreased levels of zonulin and claudin-2 proteins in the mucosal lining. These effects were notably more pronounced when compared to treatments administered individually. This study unveils a promising alternative therapy that involves combining postbiotics and 1,25D3 for treating chemotherapy-induced gut-derived P. aeruginosa sepsis.

Transcriptomics analysis highlights potential ways in human pathogenesis in Leishmania braziliensis infected with the viral endosymbiont LRV1.

The parasite Leishmania (Viannia) braziliensis is widely distributed in Brazil and is one of the main species associated with human cases of different forms of tegumentary leishmaniasis (TL) such as cutaneous leishmaniasis (CL) and mucosal leishmaniasis (ML). The mechanisms underlying the pathogenesis of TL are still not fully understood, but it is known that factors related to the host and the parasite act in a synergistic and relevant way to direct the response to the infection. In the host, macrophages have a central connection with the parasite and play a fundamental role in the defense of the organism due to their ability to destroy intracellular parasites and present antigens. In the parasite, some intrinsic factors related to the species or even the strain analyzed are fundamental for the outcome of the disease. One of them is the presence of Leishmania RNA Virus 1 (LRV1), an endosymbiont virus that parasitizes some species of Leishmania that triggers a cascade of signals leading to a more severe TL phenotype, such as ML. One of the strategies for understanding factors associated with the immune response generated after Leishmania/host interaction is through the analysis of molecular patterns after infection. Thus, the gene expression profile in human monocyte-derived macrophages obtained from healthy donors infected in vitro with L. braziliensis positive (LbLRV1+) and negative (LbLRV1-) for LRV1 was evaluated. For this, the microarray assay was used and 162 differentially expressed genes were identified in the comparison LbLRV1+ vs. LbLRV1-, 126 upregulated genes for the type I and II interferons (IFN) signaling pathway, oligoadenylate synthase OAS/RNAse L, non-genomic actions of vitamin D3 and RIG-I type receptors, and 36 down-regulated. The top 10 downregulated genes along with the top 10 upregulated genes were considered for analysis. Type I interferon (IFNI)- and OAS-related pathways results were validated by RT-qPCR and Th1/Th2/Th17 cytokines were analyzed by Cytometric Bead Array (CBA) and enzyme-linked immunosorbent assay (ELISA). The microarray results validated by RT-qPCR showed differential expression of genes related to IFNI-mediated pathways with overexpression of different genes in cells infected with LbLRV1+ compared to LbLRV1- and to the control. No significant differences were found in cytokine levels between LbLRV1+ vs. LbLRV1- and control. The data suggest the activation of gene signaling pathways associated with the presence of LRV1 has not yet been reported so far. This study demonstrates, for the first time, the activation of the OAS/RNase L signaling pathway and the non-genomic actions of vitamin D3 when comparing infections with LbLRV1+ versus LbLRV1- and the control. This finding emphasizes the role of LRV1 in directing the host's immune response after infection, underlining the importance of identifying LRV1 in patients with TL to assess disease progression.

Differences in Vitamin D3 Levels in Pulmonary Tuberculosis Patients Before and After One Month of Anti-Tuberculosis Drug Therapy

Pulmonary tuberculosis is an infectious disease caused by infection with the bacteria Mycobacterium tuberculosis. Patients diagnosed with pulmonary tuberculosis will receive Anti-Tuberculosis Drug (OAT) therapy for six months. In the first two months or what is called the intensive phase, the patient will be given a combination of OAT in the form of rifampicin, isoniazid, ethambutol and pyrazinamide. Several types of anti-tuberculosis drugs have the potential to affect vitamin D3 metabolism, either by interfering with absorption, metabolism or changing the action of vitamin D3. The long-term treatment process for pulmonary tuberculosis also results in a decrease in vitamin D3 levels in the blood. This study aims to determine the difference in vitamin D3 levels in pulmonary tuberculosis sufferers before and after one month of anti-tuberculosis drug therapy. This type of research is descriptive observational with a prospective cohort design without comparison. The total sample was 16 newly diagnosed pulmonary tuberculosis sufferers at the Bangetayu, Kedungmundu and Tlogosari Wetan Health Centers, Semarang City. Vitamin D3 levels in serum samples were examined using the ELISA method. The mean vitamin D3 level before OAT therapy was 19.00 ± 10.45 ng/mL, the level decreased after OAT therapy, namely 18.51 ± 4.74 ng/mL. These results indicate that administration of anti-tuberculosis drugs may have an effect on vitamin levels.

Hypoparathyroidism: changes in brain structure, cognitive impairment, and reduced quality of life.

Hypoparathyroidism (HypoPT) is a disease with no/or inadequate production/secretion of parathyroid hormone (PTH) from the parathyroid glands. Low levels of PTH result in hypocalcemia, which is often treated with calcium supplementation and active vitamin-D analogs. However, increasing evidence suggests that HypoPT has a profound impact on several organ systems. Quality of life (QOL) is reduced in patients with HypoPT, partly due to symptoms related to the central nervous system-including subjective feelings of confusion, a reduced ability to focus and think clearly (ie, "brain fog"). However, the extent to which these complex symptoms relate to quantifiable changes in patients' cognitive performance as determined by neuropsychological tests remains unclear. The brains of HypoPT patients may reveal tissue calcifications, but the extent to which long-term brain exposure to low PTH levels and/or changing calcium levels affects brain structure is unknown. In a cross-sectional study, we investigated PTH levels, QOL, cognitive impairment, and brain structure in well-treated post-surgical and non-surgical hypoparathyroid patients compared with healthy controls. QOL was quantified by the SF36v2, WHO-5 wellbeing Index, and two disease-specific questionnaires-the HPQ28 and Hypoparathyroidism Symptom Diary. Cognitive functions were tested using comprehensive neuropsychological. Brain structure was quantified by morphological analyses of magnetic resonance imaging images. We found reduced QOL and cognitive functioning in terms of processing speed, executive functions, visual memory, and auditory memory in HypoPT. Furthermore, HypoPT revealed a reduced volume of the hippocampus-and the size of the thalamus in postsurgical patients was associated with the disease duration. Importantly, patients reporting severe brain fog had a smaller hippocampus than those with less brainfog. HypoPT is associated with quantifiable cognitive deficits and changes in brain structure that align with patient symptoms. Our exploratory study warrants further studies of the neurobiological impact of PTH and of the impact of PTH replacements therapy on patients' cognitive functioning.

Synergistic action of vitamin D3 and A on motor activity regulation in mice model of extrapyramidal syndrome: Correlational insights into astrocyte regulation, cytokine modulation, and dopaminergic activity

BackgroundExtrapyramidal syndromes (EPS) represent neurological side effects of antipsychotic medications, characterized by motor disturbances. While previous studies have indicated the neuroprotective effects of vitamin D and A against EPS, the underlying mechanisms of this protection remain unclear. MethodsTwenty-four adult mice were categorized into four groups: positive and negative control groups, one receiving a dopamine antagonist, and the other receiving both a dopamine antagonist and vitamins D and A. Sections of the corticobasal loop, specifically the motor cortex (M1) and basal nuclei (CPu), were prepared for Immunohistochemistry (IHC) and stained with Glial Fibrillary Acidic Protein (GFAP) to visualize reactive astrocytes. ELISA assays for TNF-α, IL-6, IL-4, IL-13, and dopamine levels were performed on homogenized brain sections. ResultsThe EPS group exhibited a significant increase in TNF-α and IL-6 levels in M1 and CPu. Treatment with dopamine agonists and vitamin D&A resulted in significant reductions in IL-6 levels. Only the Vitamin D&A group showed a significant decline in TNF-α. The EPS group recorded significant decreases in IL-4 and IL-13, with IL-13 significantly elevated in the dopamine agonist and Vitamin D&A groups. IL-4 was notably increased in the Vitamin D&A groups. Dopamine concentration significantly declined in the EPS group, with improvements observed in the groups treated with dopamine agonists, and vitamin D&A. Reactive astrocytes were significantly expressed in the M1 and CPu of the EPS group but poorly expressed in other groups. ConclusionsEPS is linked to astrocyte activation, an upsurge in pro-inflammatory cytokines, a decline in anti-inflammatory cytokines, and dopamine in the corticobasal loop. Administration of vitamin D3 and A was found to suppres pro-inflammatory cytokines and repress anti-inflammatory cytokines associated with astrocyte activation.

Abstract 3867: Identification of novel biomarkers of immune toxicity from CAR T-cell therapy using ultrasensitive NULISA™ proteome technology

Abstract Introduction: The efficacy of anti-CD19 chimeric antigen receptor (CAR) T-cell therapy in patients with relapsed/refractory large B-cell lymphoma (LBCL) is limited by acute toxic events, most notably cytokine release syndrome (CRS) and immune effector cell-associated neurotoxicity syndrome (ICANS). Identification of biomarkers of toxicities can allow for selection of high-risk patients and elucidate targetable mechanisms for mitigation. Previous biomarker studies have been constrained by relatively narrow panels of potential mediators and limited time points. Here, we employed NULISA, a novel ultrasensitive assay capable of simultaneously quantifying 204 inflammatory proteins from a single sample, to identify temporal proteome associations with acute toxicities in anti-CD19 CAR T-cell treated patients. Methods: Baseline and post-treatment peripheral blood samples from 80 patients with LBCL who underwent anti-CD19 CAR-T cell therapy were collected within specific time intervals. Plasma samples (n=480) were analyzed with NULISA. CRS and ICANS were graded according to ASTCT consensus criteria. Patients were grouped according to their maximum CRS and ICANS scores. Differential protein abundance across the toxicity groups was assessed using linear mixed effects models fit for each protein, including a time by toxicity group interaction. Proteins showing significant associations were subjected to pathway enrichment and network analysis. Results: Severe CRS and ICANS patient groups showed proteome upregulation beginning on day 1-2 and peaking at day 6-9, followed by extensive downregulation on day 11-16. The strongest upregulated pathway associations of severe CRS and ICANS were inflammatory response, IL-17 signaling, non-genomic action of vitamin D3, regulation of leukocyte proliferation, and cellular extravasation. Downregulated pathways included anti-microbial humoral response and TNFs binding to receptors. In addition to the previously identified inflammatory proteins IL-6, IFNγ, sIL-2Rα, CXCL8, CCL2, our analysis revealed significant association of several novel immune regulators and mediators. These included upregulated Th2 cytokines, IL-17A, IL-22, GZMB, CTLA4, IFNA1;IFNA13, and downregulated S100A12, IL-12B, CCL22, BDNF and TNFSFs, revealing exquisite temporal orchestration of lymphoid and myeloid activities during CRS and ICANS development. Conclusions: This study represents the most comprehensive characterization of immune response to CAR T-cell therapy to date. Using the novel NULISA technology, we identified new proteins and functional pathways associated with CAR T-cell-induced toxicity, implicating them as potential biomarkers. These previously unidentified factors also provide a platform to further investigate the causative immune mechanisms of acute toxicities in CAR-T cell therapy. Citation Format: Riley Kirkpatrick, Joanne Beer, Manishkumar S. Patel, Akansha Jalota, Agrima Mian, Ishara S. Ariyapala, Qinyu Hao, Wei Feng, Xiao-Jun Ma, Yuling Luo, Brian T. Hill, Neetu Gupta. Identification of novel biomarkers of immune toxicity from CAR T-cell therapy using ultrasensitive NULISA™ proteome technology [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 3867.

1,25 dihydroxyvitamin D3-mediated effects on bovine innate immunity and on biofilm-forming Staphylococcus spp. isolated from cattle with mastitis

Mastitis is one the most widespread and serious diseases in dairy cattle. Recurrent and chronic infections are often attributable to certain pathogenicity mechanisms in mastitis-causing pathogens such as Staphylococcus spp. These include growing in biofilm and invading cells, both of which make it possible to resist or evade antimicrobial therapies and the host’s immune system. This study tested the effects of active vitamin D3 (i.e., calcitriol or 1,25-dihydroxyvitamin D3) on the internalization and phagocytosis of biofilm-forming Staphylococcus spp. isolated from animals with mastitis. Two established bovine cell lines were used: MAC-T (mammary epithelial cells) and BoMac (macrophages). Calcitriol (0–200 nM) did not affect the viability of MAC-T cells nor that of BoMac cells after 24 and 72 h. Concentrations of 0–100 mM for 24 h upregulated the expression of 24-hydroxylase in MAC-T cells, but did not alter that of VDR. Pre-treatment of the cells with calcitriol for 24 h decreased the internalization of S. aureus V329 into MAC-T cells (0–100 nM), and stimulated the phagocytosis of the same strain and of S. xylosus 4913 (0–10 nM). Calcitriol and two conditioned media, obtained by treating the cells with 25–200 nM of the metabolite for 24 h, were also assessed in terms of their antimicrobial and antibiofilm activity. Neither calcitriol by itself nor the conditioned media affected staphylococcal growth or biofilm formation (0–200 nM for 12 and 24 h, respectively). In contrast, the conditioned media (0–100 nM for 24 h) decreased the biomass of preformed non-aureus staphylococcal biofilms and killed the bacteria within them, without affecting metabolic activity. These effects may be mediated by reactive oxygen species and proteins with antimicrobial and/or antibiofilm activity. In short, calcitriol could make pathogens more accessible to antimicrobial therapies and enhance bacterial clearance by professional phagocytes. Moreover, it may modulate the host’s endogenous defenses in the bovine udder and help combat preformed non-aureus staphylococcal biofilms (S. chromogenes 40, S. xylosus 4913, and/or S. haemolyticus 6). The findings confirm calcitriol’s potential as an adjuvant to prevent and/or treat intramammary infections caused by Staphylococcus spp., which would in turn contribute to reducing antibiotic use on dairy farms.

Strain specific differences in vitamin D3 response: impact on gut homeostasis.

Vitamin D3 regulates a variety of biological processes irrespective of its well-known importance for calcium metabolism. Epidemiological and animal studies indicate a role in immune regulation, intestinal barrier function and microbiome diversity. Here, we analyzed the impact of different vitamin D3- containing diets on C57BL/6 and BALB/c mice, with a particular focus on gut homeostasis and also investigated effects on immune cells in vitro. Weak regulatory effects were detected on murine T cells. By trend, the active vitamin D3 metabolite 1,25-dihydroxyvitamin D3 suppressed IFN, GM-CSF and IL-10 cytokine secretion in T cells of C57BL/6 but not BALB/c mice, respectively. Using different vitamin D3-fortified diets, we found a tissue-specific enrichment of mainly CD11b+ myeloid cells but not T cells in both mouse strains e.g. in spleen and Peyer's Patches. Mucin Reg3γ and Batf expression, as well as important proteins for gut homeostasis, were significantly suppressed in the small intestine of C57BL76 but not BALB/c mice fed with a high-vitamin D3 containing diet. Differences between both mouse stains were not completely explained by differences in vitamin D3 receptor expression which was strongly expressed in epithelial cells of both strains. Finally, we analyzed gut microbiome and again an impact of vitamin D3 was detected in C57BL76 but not BALB/c. Our data suggest strain-specific differences in vitamin D3 responsiveness under steady state conditions which may have important implications when choosing a murine disease model to study vitamin D3 effects.

Enhancing Vitamin D3 Efficacy: Insights from Complexation with Cyclodextrin Nanosponges and Its Impact on Gut-Brain Axes in Physiology and IBS Syndrome.

Vitamin D3 (VitD3) plays a crucial role in various cellular functions through its receptor interaction. The biological activity of Vitamin D3 can vary based on its solubility and stability. Thus, the challenge lies in maximizing its biological effects through its complexation within cyclodextrin (βNS-CDI 1:4) nanosponges (NS) (defined as VitD3NS). Therefore, its activity has been evaluated on two different gut-brain axes (healthy gut/degenerative brain and inflammatory bowel syndrome gut/degenerative brain axis). At the gut level, VitD3-NS mitigated liposaccharide-induced damage (100 ng/mL; for 48 h), restoring viability, integrity, and activity of tight junctions and reducing ROS production, lipid peroxidation, and cytokines levels. Following intestinal transit, VitD3-NS improved the neurodegenerative condition in the healthy axis and the IBS model, suggesting the ability of VitD3-NS to preserve efficacy and beneficial effects even in IBS conditions. In conclusion, this study demonstrates the ability of this novel form of VitD3, named VitD3-NS, to act on the gut-brain axis in healthy and damaged conditions, emphasizing enhanced biological activity through VitD3 complexation, as such complexation increases the beneficial effect of vitamin D3 in both the gut and brain by about 50%.

P1200 Study of correlation between polymorphisms of vitamin D metabolism genes and perianal disease in Crohn's disease

Abstract Background Vitamin D (VD) is a fat-soluble vitamin essential for calcium homeostasis and that acts at the extraskeletal level. UVB skin exposure allows the synthesis of provitamin D. This undergoes a first hydroxylation in the leaver by the CYP2R1 enzyme and a second hydroxylation in the kidney by the CYP27B1 enzyme: active VD is obtained. VD is transported into the circulation by VDBP and exerts its activity in the target cells binding its VDR receptor. Finally, VD is inactivated by the renal enzyme CYP24A1. Perianal disease (pCD) is a severe phenotypic manifestation of CD that may present as perianal fistula, abscess, recto-vaginal fistula, or stenosis. Among the mechanisms involved in its pathogenesis we recognise local inflammation and intestinal microbiota alteration. Vitamin D (VD) seems to act on these elements. As there are currently no studies on this subject in the literature, the aim of this study is to evaluate the presence of an association between SNPs of genes coding for enzymes, transporters and receptors involved in the VD pathway and the occurrence of pCD in CD patients. Methods The study was carried out on the biological samples of 206 patients with CD, including 34 with pCD, who were followed up at the inflammatory bowel disease clinic in Turin, Italy. Through the use of Real-Time PCR, the genotype distribution of the following genes were assessed: VDR, CYP27B1, CYP24A1, and GC. For the association study, chi-square test was performed with calculation of p value and, when significant, logistic regression and calculation of OR with 95% CI was performed. Results Studying the association between SNPs and the presence of pCD, the following results were obtained: BsmI p=0.0470 and Apal p=0.0251. For BsmI heterozygous genotype there was an OR=2.5 (95% CI 1.2-5.3) of developing perianal disease and p value=0.02, while for ApaI heterozygous there was OR=2.91 (95% CI 1.3-6.6) of presenting pCD, with p value=0.01. Conclusion In the literature, there are several studies examining the association between the heterozygous Aa genotypes of ApaI and Bb genotypes of BsmI and increased inflammatory markers. In addition, some studies suggest that these two genotypes represent a risk factor for some diseases, including multiple myeloma, systemic lupus erythematosus, and mild cognitive disorder. This study demonstrates for the first time an impact of polymorphisms of genes associated with the VD pathway in predicting the onset of pCD. Specifically, the presence of the heterozygous genotype of BsmI and ApaI significantly increases the risk. Future studies need to be performed in different and larger cohorts of patients in order to confirm these data.

Active vitamin D increases myogenic differentiation in C2C12 cells via a vitamin D response element on the myogenin promoter.

Background: Skeletal muscle development during embryogenesis depends on proliferation of myoblasts followed by differentiation into myotubes/multinucleated myofibers. Vitamin D (VD) has been shown to affect these processes, but there is conflicting evidence within the current literature on the exact nature of these effects due to a lack of time course data. With 20%-40% of pregnant women worldwide being VD deficient, it is crucial that a clearer understanding of the impact of VD on myogenesis is gained. Methods: A detailed 8-day differentiation time course was used where C2C12 cells were differentiated in control media (2% horse serum) or with different concentrations of active VD, 1,25 (OH)2D3 (10-13M, 10-11M, 10-9M or 10-7M), and measurements were taken at 6 time points. DNA, creatine kinase and protein assays were carried out as well as quantitative PCR to determine expression of Myf5, MyoD, myogenin, MHC I, and MHC neonatal, MHC embryonic, MHC IIa, MHC IIx, and MHC IIb mRNAs. Transfections were carried out using one vector containing the myogenin promoter and another containing the same promoter with a 3 base mutation within a putative vitamin D response element (VDRE) to determine effects of 1,25 (OH)2D3 on myogenin transcription. Finally, a ChIP assay was performed to determine whether the VD receptor (VDR) binds to the putative VDRE. Results: 1,25(OH)2D3 caused an inhibition of proliferation and an increase in differentiation in C2C12 cells. Myf5, myogenin, MHC I, and MHC neonatal, MHC embryonic, MHC IIa, MHC IIx, and MHC IIb expression were all increased by 1,25(OH)2D3. Myotube size was also increased by VD. When the putative VDRE on the myogenin promoter was mutated, the increase in expression by VD was lost. ChIP analysis revealed that the VDR does bind to the putative VDRE on the myogenin promoter. Conclusion: Active VD directly increases myogenin transcription via a functional VDRE on the myogenin promoter, resulting in increased myogenic differentiation, increased expression of both the early and late MHC isoforms, and also increased myotube size. These results highlight the importance of VD status during pregnancy for normal myogenesis to occur, but further in vivo work is needed.

Vitamin D3 promotes gastric cancer cell autophagy by mediating p53/AMPK/mTOR signaling.

Objective: Vitamin D3 has the general properties of a lipid-soluble vitamin, but is also an active steroid hormone that can regulate the proliferation, apoptosis and differentiation of many tumor cells, and exerts anticancer activity against numerous malignancies. However, the mechanism underlying the effects of vitamin D3 on tumors is not fully understood. Here, we used network pharmacology and in vitro experimental approaches to explore the mechanism of vitamin D3 activity in the context of gastric cancer. Methods: The Targetnet, SuperPred, SwissTargetPrediction, and PharmMapper databases were screened for potential drug-related targets, while we used data from the PharmGKB, Drugbank, OMIM, DisGeNET, CTD, and GeneCards databases to identify potential targets associated with gastric cancer. Disease-drug crossover genes were obtained by constructing Venn diagrams. Gene ontology and Kyoto Encyclopedia of Genomes (KEGG) enrichment analyses of crossover genes were conducted and STRING was used to generate protein interaction networks and identify core targets. CCK-8 experiments were performed and apoptosis detected to assess the effect of vitamin D3 on gastric cancer cells. Western blotting was applied to detect p53/AMPK/mTOR signaling, as well as autophagy-, cell cycle-, and apoptosis-related proteins. Results: A total of 485 targets of vitamin D3 activity were obtained and 1200 gastric cancer disease-related targets discovered. Further, 60 potential targets for vitamin D3 in gastric cancer treatment were identified. KEGG analysis indicated that potential targets were mainly involved in the cell cycle, HIF-1 signaling, and the AMPK pathway, among other pathways. These findings were validated using cellular experiments, which demonstrated that the viability of AGS and SGC-7901 cells was impeded by vitamin D3. Further, vitamin D3 promoted apoptosis and inhibited the cell cycle in those cell lines, as well as activating the p53/AMPK/mTOR pathway, which promotes autophagy and inhibits tumor development. Conclusion: Our network pharmacological analyses provide preliminarily data supporting a role for vitamin D3 in promoting autophagy and apoptosis in gastric cancer cells, and in activating the p53/AMPK/mTOR pathway, which inhibits gastric cancer cell proliferation. Our findings demonstrate the molecular mechanism underlying the effect of vitamin D3 in cure of gastric cancer.

Insight into vitamin D3 action within the ovary-Basic and clinical aspects.

Vitamin D3 is a fat-soluble secosteroid predominantly synthesized in the skin or delivered with a diet. Nevertheless, recently it is considered more as a hormone than a vitamin due to its pleiotropic function within the organism ensured by widely distributed vitamin D receptors and metabolic enzymes. Besides the main role in calcium and phosphorus homeostasis, vitamin D3 was shown to regulate many cellular and metabolic processes in normal and cancerous tissues within the immune system, the cardiovascular system, the respiratory system and the endocrine system. The ovary is an important extraskeletal tissue of vitamin D3 action and local metabolism, indicating its role in the regulation of ovarian functions upon physiological and pathological conditions. This chapter reviews firstly the updated information about vitamin D3 metabolism and triggered intracellular pathways. Furthermore, the basic information about ovarian physiology and several aspects of vitamin D3 effects within the ovary are presented. Finally, the special attention is paid into possible mechanism of vitamin D3 action within ovarian pathologies such as premature ovarian failure, polycystic ovary syndrome, and ovarian cancer, considering its clinical application as alternative therapy.