Nanoparticles of nucleotide-free analogue of vitamin B12 formed in protein nanocarriers and their neuroprotective activity in vivo.

Recently, we have described the first supermolecular nanoentities of vitamin B12 derivative, viz. monocyano form of heptabutyl cobyrinate, unique nanoparticles with strong noncovalent intermolecular interactions, emerging optical and catalytic properties. Their nearest analogue, heptamethyl cobyrinate (ACCby), exhibits bioactivity. Here, we demonstrate the first example of the formation of nanoparticles of this nucleotide-free analogue of vitamin B12 in protein nanocarriers and neuroprotective activity in vivo of the own nanoform of the drug. The preparation and characterization of nanocarriers based on bovine serum albumin (BSA) loaded with vitamin B12 (viz. cyano- and aquacobalamins) and ACCby were performed. Nucleotide-free analogue of vitamin B12 is tightly retained by the protein structure and exists in an incorporated state in the form of nanoparticles. The effect of encapsulated drugs on the character and severity of primary generalized seizures in rats induced by the pharmacotoxicant thiosemicarbazide was studied. Cyanocobalamin and ACCby exhibited a neuroprotective effect. The best influence of the encapsulation on the effectiveness of the drugs was achieved in the case of AСCby, whose bioavailability as a neuroprotector did not change upon introduction in BSA particles, i.e., 33 % of surviving animals were observed upon ACCby administration in free form and in encapsulated state. No surviving rats were observed without the administration of drugs. Thus, BSA nanocarriers loaded by nanoparticles of nucleotide-free analogues of vitamin B12, including hydrophobic ones, can be recommended for neuroprotection and targeted delivery.

Vitamin B12 ameliorates gut epithelial injury via modulating the HIF-1 pathway and gut microbiota.

Inflammatory bowel diseases (IBDs) are immune chronic diseases characterized by recurrent episodes, resulting in continuous intestinal barrier damage and intestinal microbiota dysbiosis. Safe strategies aimed at stabilizing and reducing IBDs recurrence have been vigorously pursued. Here, we constructed a recurrent intestinal injury Drosophila model and found that vitamin B12 (VB12), an essential co-factor for organism physiological functions, could effectively protect the intestine and reduce dextran sulfate sodium-induced intestinal barrier disruption. VB12 also alleviated microbial dysbiosis in the Drosophila model and inhibited the growth of gram-negative bacteria. We demonstrated that VB12 could mitigate intestinal damage by activating the hypoxia-inducible factor-1 signaling pathway in injured conditions, which was achieved by regulating the intestinal oxidation. In addition, we also validated the protective effect of VB12 in a murine acute colitis model. In summary, we offer new insights and implications for the potential supportive role of VB12 in the management of recurrent IBDs flare-ups.

Nutritional vitamin B12 regulates RAS/MAPK-mediated cell fate decisions through one-carbon metabolism.

Vitamin B12 is an essential nutritional co-factor for the folate and methionine cycles, which together constitute one-carbon metabolism. Here, we show that dietary uptake of vitamin B12 modulates cell fate decisions controlled by the conserved RAS/MAPK signaling pathway in C. elegans. A bacterial diet rich in vitamin B12 increases vulval induction, germ cell apoptosis and oocyte differentiation. These effects are mediated by different one-carbon metabolites in a tissue-specific manner. Vitamin B12 enhances via the choline/phosphatidylcholine metabolism vulval induction by down-regulating fat biosynthesis genes and increasing H3K4 tri-methylation, which results in increased expression of RAS/MAPK target genes. Furthermore, the nucleoside metabolism and H3K4 tri-methylation positively regulate germ cell apoptosis and oocyte production. Using mammalian cells carrying different activated KRAS and BRAF alleles, we show that the effects of methionine on RAS/MAPK-regulated phenotype are conserved in mammals. Our findings suggest that the vitamin B12-dependent one-carbon metabolism is a limiting factor for diverse RAS/MAPK-induced cellular responses.

Vitamin B12 supplementation improved memory impairment following nicotine withdrawal in adolescent male rats: The role of oxidative stress, inflammatory, BDNF, GFAP, and AChE activity.

The present study aimed to assess the potential effect of vitamin B12 (Vit B12) on cognition impairment caused by nicotine (Nic) cessation in adolescent male rats. Adolescent male rats were categorized into two main groups as vehicle (normal saline, intraperitoneally), and Nic group in which received Nic (2 mg/kg) from 21 to 42 days of ages and then the Nic group were divided into three groups as withdrawal (the animals returned to regular diet without treatment), second and third groups received bupropion (20 mg/kg), and Vit B12 at three different doses including 0.5,1, and 1.5 mg/kg by oral gavage as treatments to attenuate Nic withdrawal symptoms. The last group including normal animals received the highest doses of Vit B12 just in the Nic abstinence period to compare the effect of that with vehicle. In MWM, Vit B12and bupropion increased the time spent in the target quadrant that is strongly associated with spatial memory as well as the more time spent with the NORT. Vit B12 and bupropion modulated both oxidant/antioxidant and inflammatory/anti-inflammatory balance, alongside inhibitory effect on AChE, and GFAP. However, BDNF and amyloid-B showed insignificant difference as compared to Vit B12 and bupropion. Considering the present results and similar related studies, Vit B12 can be introduced as a strong anti-oxidant, and anti-inflammatory agent by which probably improved memory impairment caused by Nic addiction accompanied by withdrawal. Further, other mechanisms including activity reduction of AChE, and GFAP should be considered; however, it needs further investigation and larger-scale evidences.

Evaluation of the Regeneration Effects of Systemically Applied Methylprednisolone and Vitamin B12 on Mental Nerve Crush Injury.

AIM: To evaluate the effects of methylprednisolone and vitamin B12 injection on the regeneration of the nerves after a mental nerve crush injury. MATERIAL AND METHODS: A total of 40 albino Wistar rats have been randomly divided into four groups: group 1 (n=10): crushlike nerve damage was created by pinching the left mental nerve for 60 s with an aneurysm clip and intraperitoneally administered 1 ml of saline for 14 days; group 2 (n=10): the left mental nerve was pinched for 60 s with an aneurysm clip and intraperitoneally administered 2 mg/kg of methylprednisolone for 14 days; group 3, experimental group (n=10): the left mental nerve was pinched for 60 s with an aneurysm clip and intraperitoneally administered 2 mg/kg of vitamin B12 for 14 days; and group 4, experimental group (n=10): the left mental nerve was pinched for 60 s and intraperitoneally administered 2 mg/kg of methylprednisolone and 2 mg/kg of vitamin B12 for 14 days. All rats were sacrificed on the 28th postoperative day, and histopathological evaluation was performed. RESULTS: Nerve damage was higher in the control group than in other groups (p < 0.05). When Neural Cell Adhesion Molecule (NCAM) expression levels were compared, no major differences were observed between the methylprednisolone and control groups (p > 0.05). The B12 and B12+methylprednisolone groups reached significantly higher NCAM expression levels compared to the control and methylprednisolone groups. When the myelin basic protein (MBP) expression levels were compared (p < 0.05), the MBP expression was significantly higher in all experimental groups than in the control group (p < 0.05). CONCLUSION: Systemic vitamin B12 and methylprednisolone administration effectively supported remyelination in the crushed mental nerve by increasing Schwann cell proliferation and differentiation.

Mutations Affecting Cellular Levels of Cobalamin (Vitamin B12) Confer Tolerance to Bactericidal Antibiotics in Burkholderia cenocepacia.

The Burkholderia cepacia complex (Bcc) consists of opportunistic pathogens known to cause pneumonia in immunocompromised individuals, especially those with cystic fibrosis. Treating Bcc pneumonia is challenging due to the pathogens' high multidrug resistance. Therefore, inhalation therapy with tobramycin powder, which can achieve high antibiotic concentrations in the lungs, is a promising treatment option. In this study, we investigated potential mechanisms that could compromise the effectiveness of tobramycin therapy. By selecting for B. cenocepacia survivors against tobramycin, we identified three spontaneous mutations that disrupt a gene encoding a key enzyme in the biosynthesis of cobalamin (Vitamin B12). This disruption may affect the production of succinyl-CoA by methylmalonyl-CoA mutase, which requires adenosylcobalamin as a cofactor. The depletion of cellular succinyl-CoA may impact the tricarboxylic acid (TCA) cycle, which becomes metabolically overloaded upon exposure to tobramycin. Consequently, the mutants exhibited significantly reduced reactive oxygen species (ROS) production. Both the wild-type and mutants showed tolerance to tobramycin and various other bactericidal antibiotics under microaerobic conditions. This suggests that compromised ROS-mediated killing, due to the impacted TCA cycle, underlies the mutants' tolerance to bactericidal antibiotics. The importance of ROS-mediated killing and the potential emergence of mutants that evade it through the depletion of cobalamin (Vitamin B12) provide valuable insights for developing strategies to enhance antibiotic treatments of Bcc pneumonia.

idh-1 neomorphic mutation confers sensitivity to vitamin B12 in Caenorhabditis elegans.

In humans, a neomorphic isocitrate dehydrogenase mutation (idh-1neo) causes increased levels of cellular D-2-hydroxyglutarate (D-2HG), a proposed oncometabolite. However, the physiological effects of increased D-2HG and whether additional metabolic changes occur in the presence of an idh-1neo mutation are not well understood. We created a Caenorhabditis elegans model to study the effects of the idh-1neo mutation in a whole animal. Comparing the phenotypes exhibited by the idh-1neo to ∆dhgd-1 (D-2HG dehydrogenase) mutant animals, which also accumulate D-2HG, we identified a specific vitamin B12 diet-dependent vulnerability in idh-1neo mutant animals that leads to increased embryonic lethality. Through a genetic screen, we found that impairment of the glycine cleavage system, which generates one-carbon donor units, exacerbates this phenotype. In addition, supplementation with alternate sources of one-carbon donors suppresses the lethal phenotype. Our results indicate that the idh-1neo mutation imposes a heightened dependency on the one-carbon pool and provides a further understanding of how this oncogenic mutation rewires cellular metabolism.

Propionate prevents loss of the PDIM virulence lipid in Mycobacterium tuberculosis.

Phthiocerol dimycocerosate (PDIM) is an essential virulence lipid of Mycobacterium tuberculosis. In vitro culturing rapidly selects for spontaneous PDIM-negative mutants that have attenuated virulence and increased cell wall permeability, thus impacting the relevance of experimental findings. PDIM loss can also reduce the efficacy of the BCG Pasteur vaccine. Here we show that vancomycin susceptibility can rapidly screen for M. tuberculosis PDIM production. We find that metabolic deficiency of methylmalonyl-CoA impedes the growth of PDIM-producing bacilli, selecting for PDIM-negative variants. Supplementation with odd-chain fatty acids, cholesterol or vitamin B12 restores PDIM-positive bacterial growth. Specifically, we show that propionate supplementation enhances PDIM-producing bacterial growth and selects against PDIM-negative mutants, analogous to in vivo conditions. Our study provides a simple approach to screen for and maintain PDIM production, and reveals how discrepancies between the host and in vitro nutrient environments can attenuate bacterial pathogenicity.

Disrupted one-carbon metabolism in heifers negatively affects their health and physiology.

The objective of this study was to determine the dose-dependent response of one-carbon metabolite (OCM: methionine, choline, folate, and vitamin B12) supplementation on heifer dry matter intake on fixed gain, organ mass, hematology, cytokine concentration, pancreatic and jejunal enzyme activity, and muscle hydrogen peroxide production. Angus heifers (n = 30; body weight [BW] = 392.6 ±â€…12.6 kg) were individually fed and assigned to one of five treatments: 0XNEG: total mixed ration (TMR) and saline injections at days 0 and 7 of the estrous cycle, 0XPOS: TMR, rumen-protected methionine (MET) fed at 0.08% of the diet dry matter, rumen-protected choline (CHOL) fed at 60 g/d, and saline injections at days 0 and 7, 0.5X: TMR, MET, CHOL, 5-mg B12, and 80-mg folate injections at days 0 and 7, 1X: TMR, MET CHOL, 10-mg vitamin B12, and 160-mg folate at days 0 and 7, and 2X: TMR, MET, CHOL, 20-mg vitamin B12, and 320-mg folate at days 0 and 7. All heifers were estrus synchronized but not bred, and blood samples were collected on days 0, 7, and at slaughter (day 14) during which tissues were collected. By design, heifer ADG did not differ (P = 0.96). Spleen weight and uterine weight were affected cubically (P = 0.03) decreasing from 0XPOS to 0.5X. Ovarian weight decreased linearly (P < 0.01) with increasing folate and B12 injection. Hemoglobin and hematocrit percentage were decreased (P < 0.01) in the 0.5X treatment compared with all other treatments. Plasma glucose, histotroph protein, and pancreatic α-amylase were decreased (P ≤ 0.04) in the 0.5X treatment. Heifers on the 2X treatment had greater pancreatic α-amylase compared with 0XNEG and 0.5X treatment. Interleukin-6 in plasma tended (P = 0.08) to be greater in the 0XPOS heifers compared with all other treatments. Lastly, 0XPOS-treated heifers had reduced (P ≤ 0.07) hydrogen peroxide production in muscle compared with 0XNEG heifers. These data imply that while certain doses of OCM do not improve whole animal physiology, OCM supplementation doses that disrupt one-carbon metabolism, such as that of the 0.5X treatment, can induce a negative systemic response that results in negative effects in both the dam and the conceptus during early gestation. Therefore, it is necessary to simultaneously establish an optimal OCM dose that increases circulating concentrations for use by the dam and the conceptus, while avoiding potential negative side effects of a disruptive OCM, to evaluate the long-term impacts of OCM supplementation of offspring programming.

The feeding of one-carbon metabolites (including methionine and B vitamins) has been shown to improve fetal growth and milk production in species such as mice, sheep, and dairy cattle. Extending this to beef cattle around the time of breeding is a growing area of research. Our group previously determined that one-carbon metabolite supplementation to beef heifers altered the abundance of circulating methionine-folate cycle intermediates in a dose-dependent manner. Therefore, we aimed to determine a whole-body response to one-carbon metabolite supplementation in heifers by measuring the effects on specific physiological systems as well as a total systemic response. We determined that treatments that negatively altered the methionine-folate cycle yielded a fundamental negative whole-body response to supplementation.

Elucidating the structural basis of vitamin B12 derivatives as novel potent inhibitors of PTP1B: Insights from inhibitory mechanisms using Gaussian accelerated molecular dynamics (GaMD) and in vitro study.

Vitamin B12 is a group of biologically active cobalamin compounds. In this study, we investigated the inhibitory effects of methylcobalamin (MeCbl) and hydroxocobalamin acetate (OHCbl Acetate) on protein tyrosine phosphatase 1B (PTP1B). MeCbl and OHCbl Acetate exhibited an IC50 of approximately 58.390 ± 2.811 µM and 8.998 ± 0.587 µM, respectively. The Ki values of MeCbl and OHCbl Acetate were 25.01 µM and 4.04 µM respectively. To elucidate the inhibition mechanism, we conducted a 500 ns Gaussian accelerated molecular dynamics (GaMD) simulation. Utilizing PCA and tICA, we constructed Markov state models (MSM) to examine secondary structure changes during motion. Our findings revealed that the α-helix at residues 37-42 remained the most stable in the PTP1B-OHCbl Acetate system. Furthermore, upon binding of OHCbl Acetate or MeCbl, the WPD loop of PTP1B moved inward to the active pocket, forming a closed conformation and potentially obstructs substrate entry. Protein-ligand interaction analysis and MM-PBSA showed that OHCbl Acetate exhibited lower binding free energy and engaged in more residue interactions with PTP1B. In summary, our study confirmed the substantial inhibitory activity of OHCbl Acetate against PTP1B, with its inhibitory potency notably surpassing that of MeCbl. We demonstrated potential molecular mechanisms of OHCbl Acetate inhibiting PTP1B.

Cobalamins Function as Allosteric Activators of an Angelman Syndrome-Associated UBE3A/E6AP Variant.

Genetic aberrations of the maternal UBE3A allele, which encodes the E3 ubiquitin ligase E6AP, are the cause of Angelman syndrome (AS), an imprinting disorder. In most cases, the maternal UBE3A allele is not expressed. Yet, approximately 10 percent of AS individuals harbor distinct point mutations in the maternal allele resulting in the expression of full-length E6AP variants that frequently display compromised ligase activity. In a high-throughput screen, we identified cyanocobalamin, a vitamin B12-derivative, and several alloxazine derivatives as activators of the AS-linked E6AP-F583S variant. Furthermore, we show by cross-linking coupled to mass spectrometry that cobalamins affect the structural dynamics of E6AP-F583S and apply limited proteolysis coupled to mass spectrometry to obtain information about the regions of E6AP that are involved in, or are affected by binding cobalamins and alloxazine derivatives. Our data suggest that dietary supplementation with vitamin B12 can be beneficial for AS individuals.

Dietary Supplementation of Vitamin B12 to Rats Fed High-Amylose Cornstarch Normalizes Propionate Fermentation in the Colon.

Since propionate exerts several physiological effects, maintenance of its normal colonic fermentation is essential. To investigate whether vitamin B12 (VB12) is essential for normal propionate fermentation by colonic bacteria, via the succinate pathway, we examined if high-amylose cornstarch (HACS) feeding activated such a pathway, if high HACS feeding impaired propionate fermentation, and if oral VB12 supplementation normalized propionate fermentation. Male rats were given control, 20% HACS or 3% fucose diets (Expt. 1); a VB12-free control diet or one supplemented with 5-30% HACS (Expt. 2); and the 20% HACS diet supplemented with 0.025-25 mg/kg of VB12 (Expt. 3), for 14 d. HACS feeding significantly increased cecal succinate concentration, activating the succinate pathway (Expt. 1). Cecal cobalamin concentration in 20% and 30% HACS groups was about 75% of that in the control group (Expt. 2). Cecal succinate and propionate concentrations significantly increased and decreased in 30% HACS groups, respectively, compared with the control group. Although HACS group supplemented with 0.025 mg/kg of VB12 had a low concentration of cecal propionate, adding high amounts of VB12 to HACS diets provided sufficient amounts of VB12 to rat ceca and increased cecal propionate concentration (Expt. 3). Compared with the non-HACS group, the relative abundance of Akkermansia muciniphila, but not Bacteroides/Phocaeicola, was lower in the HACS counterpart and showed improvement with increased VB12 doses. To summarize, feeding high HACS decreased and increased cecal VB12 and succinate concentrations, respectively. Furthermore, colonic delivery of sufficient amounts of VB12 to rats likely reduced accumulation of succinate and normalized propionate fermentation.

Rescue of Methionine Dependence by Cobalamin in a Human Colorectal Cancer Cell Line.

Methionine dependence is a characteristic of most cancer cells where they are unable to proliferate when the essential amino acid methionine is replaced with its precursor homocysteine in the growing media. Normal cells, on the other hand, thrive under these conditions and are referred to as methionine-independent. The reaction that adds a methyl group from 5-methyltetrahydrofolate to homocysteine to regenerate methionine is catalyzed by the enzyme methionine synthase with the cofactor cobalamin (vitamin B12). However, decades of research have shown that methionine dependence in cancer is not due to a defect in the activity of methionine synthase. Cobalamin metabolism has been tied to the dependent phenotype in rare cell lines. We have identified a human colorectal cancer cell line in which the cells regain the ability to proliferation in methionine-free, L-homocystine-supplemented media when cyanocobalamin is supplemented at a level of 1 µg/mL. In human SW48 cells, methionine replacement with L-homocystine does not induce any measurable increase in apoptosis or reactive oxygen species production in this cell line. Rather, proliferation is halted, then restored in the presence of cyanocobalamin. Our data show that supplementation with cyanocobalamin prevents the activation of the integrated stress response (ISR) in methionine-deprived media in this cell line. The ISR-associated cell cycle arrest, characteristic of methionine-dependence in cancer, is also prevented, leading to the continuation of proliferation in methionine-deprived SW48 cells with cobalamin. Our results highlight differences between cancer cell lines in the response to cobalamin supplementation in the context of methionine dependence.

Vitamin B12 Protects against Genotoxicity Induced by Cisplatin.

BACKGROUND: Cisplatin is an effective synthetic chemotherapeutic drug used for cancer treatment. Vitamin B12 has been shown to possess anti-genotoxic activity. This study aimed to investigate the effect of vitamin B12 on chromosomal damage induced by cisplatin. METHODS: The level of sister chromatid exchanges (SCEs) and chromosomal aberrations (CAs) were measured in cultured human blood lymphocytes treated with cisplatin and/or vitamin B12. RESULTS: The results showed a significantly elevated frequency of CAs and SCEs of cisplatin-treated cultures compared to the control (P < 0.05). The CAs and SCEs induced by cisplatin were significantly lowered by pretreatment of cell cultures with vitamin B12. In addition, cisplatin caused a slight reduction in the mitotic index (MI), while vitamin B12 did not modulate the effect of cisplatin on MI. CONCLUSION: Vitamin B12 can protect human lymphocytes against genotoxicity associated with cisplatin.

One-carbon metabolite supplementation to nutrient-restricted beef heifers affects placental vascularity during early pregnancy.

We hypothesized that restricted maternal nutrition and supplementation of one-carbon metabolites (OCM; methionine, folate, choline, and vitamin B12) would affect placental vascular development during early pregnancy. A total of 43 cows were bred, and 32 heifers successfully became pregnant with female calves, leading to the formation of four treatment groups: CON - OCM (n = 8), CON + OCM (n = 7), RES - OCM (n = 9), and RES + OCM (n = 8). The experimental design was a 2 × 2 factorial, with main factors of dietary intake affecting average daily gain: control (CON; 0.6 kg/d ADG) and restricted (RES; -0.23 kg/d ADG); and OCM supplementation (+OCM) in which the heifers were supplemented with rumen-protected methionine (7.4 g/d) and choline (44.4 g/d) and received weekly injections of 320 mg of folate and 20 mg of vitamin B12, or received no supplementation (-OCM; corn carrier and saline injections). Heifers were individually fed and randomly assigned to treatment at breeding (day 0). Placentomes were collected on day 63 of gestation (0.225 of gestation). Fluorescent staining with CD31 and CD34 combined with image analysis was used to determine the vascularity of the placenta. Images were analyzed for capillary area density (CAD) and capillary number density (CND). Areas evaluated included fetal placental cotyledon (COT), maternal placental caruncle (CAR), whole placentome (CAR + COT), intercotyledonary fetal membranes (ICOT, or chorioallantois), intercaruncular endometrium (ICAR), and endometrial glands (EG). Data were analyzed with the GLM procedure of SAS, with heifer as the experimental unit and significance at P ≤ 0.05 and a tendency at P > 0.05 and P < 0.10. Though no gain × OCM interactions existed (P ≥ 0.10), OCM supplementation increased (P = 0.01) CAD of EG, whereas nutrient restriction tended (P < 0.10) to increase CAD of ICOT and CND of COT. Additionally, there was a gain × OCM interaction (P < 0.05) for CAD within the placentome and ICAR, such that RES reduced and supplementation of RES with OCM restored CAD. These results indicate that maternal rate of gain and OCM supplementation affected placental vascularization (capillary area and number density), which could affect placental function and thus the efficiency of nutrient transfer to the fetus during early gestation.

In cow­calf production, periods of poor forage availability or quality can result in nutrient restriction during pregnancy. Previous studies have shown that even moderate maternal feed restriction during pregnancy, including very early in pregnancy, has profound effects on fetal and placental development, potentially having lasting impacts on calf growth and body composition later in life. One-carbon metabolites (OCM) in the diet are biomolecules required for methylation reactions and participate in the regulation of gene expression. Our objective was to evaluate the effects of nutrient restriction and OCM supplementation (specifically methionine, choline, folate, and vitamin B12) on placental vascular development during early pregnancy. Proper placental vascular development is necessary for healthy pregnancy outcomes, reflected by normal birth weight and healthy offspring. Our results indicated that maternal rate of gain and OCM supplementation affect placental vascularization, which could affect placental function and thereby fetal development throughout gestation. In the context of beef cattle production, our study sheds light on strategies that could enhance placental vascular development during early pregnancy. However, it is essential to recognize the nuances in our data, highlighting the need for further research to fully comprehend these intricate processes.

Effects of biotin and coated cobalamin on lactation performance, nutrient digestion and rumen fermentation in Holstein dairy cows.

Biotin (BI) and cobalamin (CA) are essential for rumen propionate production and hepatic gluconeogenesis. The study evaluated the influence of BI or/and coated CA (CCA) on milk performance and nutrient digestion in cows. Sixty Holstein dairy cows were assigned in a 2 × 2 factorial arrangement and randomised block design to four groups. The factors were BI at 0 or 20 mg/day and CCA at 0 or 9 mg CA/day. Dry matter intake increased with BI addition but was unchanged with CCA supply. Addition of BI or CCA increased fat-corrected milk, milk fat and milk protein yields and feed efficiency. Moreover, lactose yield was increased by CCA addition. Dry matter, organic matter, crude protein and acid detergent fibre total-tract digestibility increased for BI or CCA supply. When CCA was supplemented, positive response of neutral detergent fibre digestibility to BI addition was enhanced. Supplementing BI did not affect pH, propionate content and acetate to propionate ratio, but increased total volatile fatty acids (VFA) and acetate contents. Supplementing CCA decreased pH and acetate to propionate ratio, but increased total VFA, acetate and propionate contents. Rumen protease and carboxymethyl-cellulase activities and fungi, bacteria and Butyrivibrio fibrisolvens numbers increased for BI or CCA supply. In addition, protozoa increased for BI addition, and protease activity and Prevotella ruminicola increased for CCA supply. When CCA was supplemented, positive responses of R. albus and Ruminobacter amylophilus numbers to BI addition were enhanced. Blood glucose concentration was unchanged with BI supply, but increased for CCA supply. Blood nonesterified fatty acids and ß-hydroxybutyrate contents reduced with BI or CCA supply. Supplementation with BI or CCA increased blood BI or CA content. The results showed that supplementing BI or/and CCA improved lactation performance and nutrient digestion, and CCA supply did not enhance the lactation performance response to BI supply.

Subacute Combined Degeneration of the Spinal Cord Induced by Nitrous Oxide Abuse: A Rare Patient Presentation to a Spine Surgery Clinic: Illustrative Case.

Subacute combined degeneration (SCD) of the spinal cord is a disease involving the lateral and posterior columns of the spinal cord that can manifest in patients with vitamin B12 deficiency. Nitrous oxide (N2O)-induced SCD of the spinal cord is a result of N2O interfering with the metabolism of vitamin B12 and results in nervous system demyelination. This is an infrequent complication of N2O anesthesia; however, cases are rising with recreational N2O use. This case report describes a patient with SCD of the spinal cord induced by recreational N2O abuse. The patient presented to a spine surgery clinic with a 3-week history of progressive global weakness and paresthesias. After a detailed history and physical examination, the diagnosis was made and supported by various tests and imaging findings. Despite marked neurologic deficits, the patient's symptoms improved markedly with therapy and vitamin B12 supplementation. Spine surgery clinicians may be confronted with these cases and should be aware of this atypical presentation of SCD. As in our case, patients may present with neurologic deficits of unclear etiology. Neurologic dysfunction may be irreversible; therefore, accurate diagnosis, medical treatment, and complete neurologic evaluation are of the utmost importance to prevent additional progression.

Impact of vitamin B12 on rhamnose metabolism, stress defense and in-vitro virulence of Listeria monocytogenes.

Listeria monocytogenes is a facultative anaerobe which can cause a severe food-borne infection known as listeriosis. L. monocytogenes is capable of utilizing various nutrient sources including rhamnose, a naturally occurring deoxy sugar abundant in foods. L. monocytogenes can degrade rhamnose into lactate, acetate and 1,2-propanediol. Our previous study showed that addition of vitamin B12 stimulated anaerobic growth of L. monocytogenes on rhamnose due to the activation of bacterial microcompartments for 1,2-propanediol utilization (pdu BMC) with concomitant production of propionate and propanol. Notably, anaerobic 1,2-propanediol metabolism has been linked to virulence of enteric pathogens including Salmonella spp. and L. monocytogenes. In this study we investigated the impact of B12 and BMC activation on i) aerobic and anerobic growth of L. monocytogenes on rhamnose and ii) the level of virulence. We observed B12-induced pdu BMC activation and growth stimulation only in anaerobically grown cells. Comparative Caco-2 virulence assays showed that these pdu BMC-induced cells have significantly higher translocation efficiency compared to non-induced cells (anaerobic growth without B12; aerobic growth with or without B12), while adhesion and invasion capacity is similar for all cells. Comparative proteome analysis showed specific and overlapping responses linked to metabolic shifts, activation of stress defense proteins and virulence factors, with RNA polymerase sigma factor SigL, teichoic acid export ATP-binding protein TagH, DNA repair and protection proteins, RadA and DPS, and glutathione synthase GshAB, previously linked to activation of virulence response in L. monocytogenes, uniquely upregulated in anaerobically rhamnose grown pdu-induced cells. Our results shed light on possible effects of B12 on L. monocytogenes competitive fitness and virulence activation when utilizing rhamnose in anaerobic conditions encountered during transmission and the human intestine.

Neuroprotective role of vitamin B12 in streptozotocin-induced type 1 diabetic rats.

Chronic hyperglycemia-induced neuropathological changes include neuronal apoptosis, astrogliosis, decrease in neurotrophic support, impaired synaptic plasticity, and impaired protein quality control (PQC) system. Vitamin B12 is indispensable for neuronal development and brain function. Several studies reported the neuroprotective effect of B12 supplementation in diabetic patients. However, the underlying molecular basis for the neuroprotective effect of B12 supplementation in diabetes needs to be thoroughly investigated. Two-month-old Sprague-Dawley rats were randomly assigned into three groups: Control (CN), diabetes (D; induced with streptozotocin; STZ), and diabetic rats supplemented with vitamin B12 (DBS; vitamin B12; 50 µg/kg) for four months. At the end of 4 months of experimentation, the brain was dissected to collect the cerebral cortex (CC). The morphology of CC was investigated with H&E and Nissl body staining. Neuronal apoptosis was determined with TUNEL assay. The components of neurotrophic support, astrogliosis, synaptic plasticity, and PQC processes were investigated by immunoblotting and immunostaining methods. H& E, Nissl body, and TUNEL staining revealed that diabetes-induced neuronal apoptosis and degeneration. However, B12 supplementation ameliorated the diabetes-induced neuronal apoptosis. Further, B12 supplementation restored the markers of neurotrophic support (BDNF, NGF, and GDNF), and synaptic plasticity (SYP, and PSD-95) in diabetic rats. Interestingly, B12 supplementation also attenuated astrogliosis, ER stress, and ameliorated autophagy-related proteins in diabetic rats. Overall, these findings suggest that B12 acts as a neuroprotective agent by inhibiting the neuropathological changes in STZ-induced type 1 diabetes. Thus, B12 supplementation could produce beneficial outcomes including neuroprotective effects in diabetic patients.

Improving the quality of chilled semen from Thai native chicken using phosphorus and vitamin B12 supplementation in semen extender.

This study aimed to determine phosphorus and vitamin B12 supplementation effect in semen extender on the quality and fertility ability of chilled Thai native rooster semen. Eighty-four ejaculates of semen from 26 Thai native roosters (Burmese × Vietnam crossbreed) were included. Semen was collected by applying dorsal-abdominal massage once a week, pooled, diluted to 500 million sperms per dose, and divided into 6 groups. The semen samples used for control group were diluted with modified Beltsville poultry semen extender (BPSE). For the treatment groups 2 to 6: semen samples were diluted with modified BPSE and enriched with phosphorus and vitamin B12 (Octafos Octa Memorial Co., Ltd., Bangkok, Thailand) at concentrations 0.02, 0.04, 0.06, 0.08, and 0.10%. Semen fertility ability was tested in 6 replications by inseminating layer hens. Thirty-six Thai native hens were randomly assigned to 3 groups (control, 0.04, and 0.08%) of 12 hens and were inseminated with a dose of 0.2 mL on collecting day. Sperm motion characteristics (i.e., sperm motility, sperm progressive motility, and sperm kinetic parameters) were measured using a computer-assisted sperm analysis system (SCA, Proiser S.L., Valencia, Spain). Sperm viability, mitochondrial activity, acrosome integrity, plasma membrane integrity, and malondialdehyde (MDA) concentration were also evaluated. The sperm motion characteristics were the highest in the 0.04% supplementation group on all days of collection, especially the VCL and VAP (P < 0.05). The viability, mitochondrial activity, plasma membrane and acrosome integrity of spermatozoa were greater in the 0.04% supplementation group than in the control groups (P < 0.05). The 0.04% supplementation group had the lowest MDA concentration in all days of collection. The 0.04% supplementation group were higher both fertility (66.59 vs. 48.50%: P < 0.05) and hatching rates (58.80 vs. 43.18%: P < 0.05) than in the control group. In conclusion, 0.04% phosphorus and vitamin B12 concentrations supplementation in semen extender improved rooster semen quality and fertility in chilled rooster semen.