Thiamin deficiency induces impaired fish gill immune responses, tight junction protein expression and antioxidant capacity: Roles of the NF-κB, TOR, p38 MAPK and Nrf2 signaling molecules.

In this study, we investigate the effects of dietary thiamin deficiency on immune responses, tight junctions, antioxidant capacity and related signaling molecules in the gills of young grass carp (Ctenopharyngodon idella). Fish were fed diets that contained 0.12-2.04 mg thiamin kg(-1) for 8 weeks. We found that dietary thiamin deficiency resulted in reduced complement 3 content, lysozyme and acid phosphatase activities, mRNA levels of hepcidin, liver-expressed antimicrobial peptides 2, transforming growth factor (TGF)-ß1, interleukin (IL)-10, inhibitor protein-κBα (IκBα), ribosomal S6 protein kinase 1 and target of rapamycin (TOR) and increased expression of interferon-γ2, tumor necrosis factor-α, TGF-ß2, IL-1ß, IL-8, IκB kinases (IKKß and IKKγ) and nuclear factor-κB p65 (NF-κB p65). Our findings showed that thiamin deficiency reduced the immune status of fish gills. Furthermore, thiamin deficiency resulted in reduced mRNA transcript levels of claudin b, claudin 3, claudin 12, zonula occludens 1 (ZO-1) and occludin and increased mRNA transcript levels of claudin 15a, myosin light-chain kinase (MLCK) and p38 mitogen-activated protein kinase (p38 MAPK) in fish gill tissues. These data suggested that thiamin deficiency disrupted tight junction-mediated fish gill barrier function. Additionally, reactive oxygen species, malondialdehyde and protein carbonyl levels and both the activities and expression levels of Cu/Zn superoxide dismutase, catalase, glutathione peroxidase, glutathione-S-transferases and glutathione reductase, as well as NF-E2-related factor 2 gene expression in fish gills, were lower in fish fed a thiamin-deficient diet. By contrast, thiamin deficiency increased levels of Kelch-like-ECH-associated protein 1a (Keap1a) and Keap1b mRNA transcript expression in fish gills. Taken together, our findings indicated that thiamin deficiency impaired fish gill health by effects on the expression of genes encoding cytokines, tight junction proteins, antioxidant enzymes, NF-κB p65, MLCK and Nrf2.

Evaluation the effect of thiamin deficiency on intestinal immunity of young grass carp (Ctenopharyngodon idella).

Our study explored the effect of dietary thiamin on growth and immunity (intestine, head kidney, spleen and liver) of young grass carp (Ctenopharyngodon idella). Fish were fed diets containing six graded levels of thiamin at 0.12-2.04 mg/kg diet for 8 weeks. The percentage weight gain (PWG), feed intake and feed efficiency were lower in fish fed the 0.12 mg/kg diet. Thiamin deficiency decreased complement 3 content, lysozyme (LA) and acid phosphatase activities, mRNA levels of hepcidin and interleukin (IL) 10, elevated mRNA levels of interferon γ2, tumor necrosis factor α, IL-1ß and IL-8 in intestine, head kidney, spleen and liver. The mRNA levels of inhibitor protein-κBα, target of rapamycin (TOR) and NF-E2-related factor 2 (Nrf2), the activities and mRNA levels of copper/zinc superoxide dismutase, manganese superoxide dismutase, catalase, glutathione peroxidase, glutathione-S-transferase and glutathione reductase were down-regulated, mRNA levels of myosin light-chain kinase (MLCK), IκB kinases (IKKß and IKKγ), nuclear factor κB P65 (NF-κB P65) and Kelch-like-ECH-associated protein 1a (Keap1a) were up-regulated in the intestine of fish fed the thiamin-deficient diet. Additionally, thiamin deficiency decreased claudin b, c and 3, ZO-1 and occludin mRNA levels in each intestinal segment, increased claudin 12 and claudin 15a mRNA levels in distal intestine. In conclusion, thiamin deficiency decreased fish growth and immunity of intestine, head kidney, spleen and liver. The dietary thiamin requirement of young grass carp (242-742 g) based on intestinal LA activity or PWG were determined to be 1.15 or 0.90 mg/kg diet, respectively.

Thiamine deficiency promotes T cell infiltration in experimental autoimmune encephalomyelitis: the involvement of CCL2.

Multiple sclerosis (MS) is a complex multifactorial disease that results from the interplay between environmental factors and a susceptible genetic background. Experimental autoimmune encephalomyelitis (EAE) has been widely used to investigate the mechanisms underlying MS pathogenesis. Chemokines, such as CCL2, are involved in the development of EAE. We have previously shown that thiamine deficiency (TD) induced CCL2 in neurons. We hypothesized that TD may affect the pathogenesis of EAE. In this study, EAE was induced in C57BL/6J mice by the injection of myelin oligodendroglial glycoprotein (MOG) peptides 35-55 with or without TD. TD aggravated the development of EAE, which was indicated by clinical scores and pathologic alterations in the spinal cord. TD also accelerated the development of EAE in an adoptive transfer EAE model. TD caused microglial activation and a drastic increase (up 140%) in leukocyte infiltration in the spinal cord of the EAE mice; specifically, TD increased Th1 and Th17 cells. TD upregulated the expression of CCL2 and its receptor CCR2 in the spinal cord of EAE mice. Cells in peripheral lymph node and spleen isolated from MOG-primed TD mice showed much stronger proliferative responses to MOG. CCL2 stimulated the proliferation and migration of T lymphocytes in vitro. Our results suggested that TD exacerbated the development of EAE through activating CCL2 and inducing pathologic inflammation.

In vitro immune functions in thiamine-replete and -depleted lake trout (Salvelinus namaycush).

In this study we examined the impacts of in vivo thiamine deficiency on lake trout leukocyte function measured in vitro. When compared outside the context of individual-specific thiamine concentrations no significant differences were observed in leukocyte bactericidal activity or in concanavalin A (Con A), and phytohemagglutinin-P (PHA-P) stimulated leukocyte proliferation. Placing immune functions into context with the ratio of in vivo liver thiamine monophosphate (TMP--biologically inactive form) to thiamine pyrophosphate (TPP--biologically active form) proved to be the best indicator of thiamine depletion impacts as determined using regression modeling. These observed relationships indicated differential effects on the immune measures with bactericidal activity exhibiting an inverse relationship with TMP to TPP ratios, Con A stimulated mitogenesis exhibiting a positive relationship with TMP to TPP ratios and PHA-P stimulated mitogenesis exhibiting no significant relationships. In addition, these relationships showed considerable complexity which included the consistent observation of a thiamine-replete subgroup with characteristics similar to those seen in the leukocytes from thiamine-depleted fish. When considered together, our observations indicate that lake trout leukocytes experience cell-type specific impacts as well as an altered physiologic environment when confronted with a thiamine-limited state.

Impact of thiamine deficiency on T-cell dependent and T-cell independent antibody production in lake trout.

Lake trout Salvelinus namaycush on thiamine-replete and thiamine-depleted diets were evaluated for the effects of thiamine status on in vivo responses to the T-dependent antigen trinitophenol (TNP)-keyhole limpet hemocyanin (TNP-KLH), the T-independent antigen trinitrophenol-lipolysaccaharide (TNP-LPS), or Dulbecco's phosphate-buffered saline (DPBS; negative control fish). Plasma antibody concentrations were evaluated for possible differences in total anti-TNP activity as well as differences in response kinetics. Associations between anti-TNP activity and muscle and liver thiamine concentrations as well as ratios of muscle-to-liver thiamine to anti-TNP activity were also examined. Thiamine-depleted lake trout that were injected with TNP-LPS exhibited significantly more anti-TNP activity than thiamine-replete fish. The depleted fish injected with TNP-LPS also exhibited significantly different response kinetics relative to thiamine-replete lake trout. No differences in activity or kinetics were observed between the thiamine-replete and -depleted fish injected with TNP-KLH or in the DPBS negative controls. Anti-TNP activity in thiamine-depleted lake trout injected with TNP-KLH was positively associated with muscle thiamine pyrophosphate (thiamine diphosphate; TPP) concentration. A negative association was observed between the ratio of muscle-to-liver TPP and T-independent responses. No significant associations between anti-TNP activity and tissue thiamine concentration were observed in the thiamine-replete fish. We demonstrated that thiamine deficiency leads to alterations in both T-dependent and T-independent immune responses in lake trout.

Neuronal MCP-1 mediates microglia recruitment and neurodegeneration induced by the mild impairment of oxidative metabolism.

Chemokines are implicated in the neuroinflammation of several chronic neurodegenerative disorders. However, the precise role of chemokines in neurodegeneration is unknown. Thiamine deficiency (TD) causes abnormal oxidative metabolism in the brain as well as a well-defined microglia activation and neurodegeneration in the submedial thalamus nucleus (SmTN), which are common features of neurodegenerative diseases. We evaluated the role of chemokines in neurodegeneration and the underlying mechanism in a TD model. Among the chemokines examined, TD selectively induced neuronal expression of monocyte chemoattractant protein-1 (MCP-1) in the SmTN prior to microglia activation and neurodegeneration. The conditioned medium collected from TD-induced neurons caused microglia activation. With a neuron/microglia co-culture system, we showed that MCP-1-induced neurotoxicity required the presence of microglia, and exogenous MCP-1 was able to activate microglia and stimulated microglia to produce cytokines. A MCP-1 neutralizing antibody inhibited MCP-1-induced microglia activation and neuronal death in culture and in the thalamus. MCP-1 knockout mice were resistant to TD-induced neuronal death in SmTN. TD selectively induced the accumulation of reactive oxygen species in neurons, and antioxidants blocked TD-induced MCP-1 expression. Together, our results indicated an induction of neuronal MCP-1 during mild impairment of oxidative metabolism caused by microglia recruitment/activation, which exacerbated neurodegeneration.

Thiamine deficiency induces oxidative stress and exacerbates the plaque pathology in Alzheimer's mouse model.

Mitochondrial dysfunction, oxidative stress and reductions in thiamine-dependent enzymes have been implicated in multiple neurological disorders including Alzheimer's disease (AD). Experimental thiamine deficiency (TD) is an established model for reducing the activities of thiamine-dependent enzymes in brain. TD diminishes thiamine-dependent enzymes throughout the brain, but produces a time-dependent selective neuronal loss, glial activation, inflammation, abnormalities in oxidative metabolism and clusters of degenerating neurites in only specific thalamic regions. The present studies tested how TD alters brain pathology in Tg19959 transgenic mice over expressing a double mutant form of the amyloid precursor protein (APP). TD exacerbated amyloid plaque pathology in transgenic mice and enlarged the area occupied by plaques in cortex, hippocampus and thalamus by 50%, 200% and 200%, respectively. TD increased Abeta(1-42) levels by about three fold, beta-CTF (C99) levels by 33% and beta-secretase (BACE1) protein levels by 43%. TD-induced inflammation in areas of plaque formation. Thus, the induction of mild impairment of oxidative metabolism, oxidative stress and inflammation induced by TD alters metabolism of APP and/or Abeta and promotes accumulation of plaques independent of neuron loss or neuritic clusters.

Gene expression changes in thalamus and inferior colliculus associated with inflammation, cellular stress, metabolism and structural damage in thiamine deficiency.

Identification of gene expression changes that promote focal neuronal death and neurological dysfunction can further our understanding of the pathophysiology of these disease states and could lead to new pharmacological and molecular therapies. Impairment of oxidative metabolism is a pathogenetic mechanism underlying neuronal death in many chronic neurodegenerative diseases as well as in Wernicke's encephalopathy (WE), a disorder induced by thiamine deficiency (TD). To identify functional pathways that lead to neuronal damage in this disorder, we have examined gene expression changes in the vulnerable thalamus and inferior colliculus of TD rats using Affymetrix Rat Genome GeneChip analysis in combination with gene ontology and functional categorization assessment utilizing the NetAffx GO Mining Tool. Of the 15 927 transcripts analysed, 125 in thalamus and 141 in inferior colliculus were more abundantly expressed in TD rats compared with control animals. In both regions, the major functional categories of transcripts that were increased in abundance after TD were those associated with inflammation (approximately 33%), stress (approximately 20%), cell death and repair ( approximately 26%), and metabolic perturbation (approximately 19%), together constituting approximately 98% of all transcripts up-regulated. These changes occurred against a background of neuronal cell loss and reactive astro- and microgliosis in both structures. Our results indicate that (i) TD produces changes in gene expression that are consistent with the observed dysfunction and pathology, and (ii) similar alterations in expression occur in thalamus and inferior colliculus, brain regions previously considered to differ in pathology. These findings provide important new insight into processes responsible for lesion development in TD, and possibly WE.

CD40-CD40L interactions promote neuronal death in a model of neurodegeneration due to mild impairment of oxidative metabolism.

Abnormalities in oxidative processes, region-selective neuron loss, inflammation and diminished activity of thiamine-dependent enzymes characterize age-related neurodegenerative diseases. Thiamine deficiency (TD) models the selective neurodegeneration that accompanies mild impairment of oxidative metabolism. As in human neurodegenerative diseases, alterations in multiple cell types accompany the TD-induced neurodegeneration. The current studies demonstrate that CD40 and CD40 ligand (CD40L), two co-stimulatory immune molecules, are involved in TD-induced selective neuronal death. TD induced CD40 immunoreactivity in microglia and CD40L immunoreactivity in astrocytes. Both CD40-positive microglia and CD40L-positive astrocytes increased during the progressive TD-induced neuronal death. In early stages of TD, targeted deletion of CD40 diminished the number of CD40L-positive astrocytes and reduced neuronal death by 35%. The number of CD40L-positive astrocytes increased whenever the number of NeuN-positive neurons decreased. In early stages of TD, deletion of CD40L diminished CD40-positive microglia and reduced the neuronal death by 64%. In advanced phases of TD, neither CD40 nor CD40L deletion protected against neuronal death. The data show for the first time that TD induces expression of CD40 by the microglia and CD40L by astrocytes. The results indicate that CD40-CD40L interactions promote neuronal death in early stages of TD, but that at later phases the protective effects of the diminished CD40 or CD40L are over-ridden by other mechanisms.

Neuropathological study of the role of mast cells and histamine-positive neurons in selective vulnerability of the thalamus and inferior colliculus in thiamine-deficient encephalopathy.

The purpose of the present study was to examine the role of histamine in the pathogenesis of experimental thiamine-deficient encephalopathy. By studying sagittal serial sections the authors were able to examine the topographical relationship between histamine-positive neurons and fibers, the number of mast cells, and localized lesions in the thalamus (TH) and inferior colliculus (IC). Adult rats were given a thiamine-deficient diet and pyrithiamine was given intraperitoneally (30 microg/100 g bodyweight per day), and the distribution of vulnerable regions and petechial bleeding was histologically examined by reconstruction of the sagittal serial sections. The distribution of mast cells and histamine-positive neurons and fibers was examined immunohistochemically in control rats, and compared between the vulnerable and non-vulnerable regions of the TH and tectum. Changes in the aforementioned measures during the thiamine-deficient state were also examined. The blood-brain barrier was examined using antibodies against rat endothelial barrier antigen (EBA) and albumin. The density of histamine-positive fibers in the vulnerable regions of the TH and IC was very low and not different from the non-vulnerable regions, and the number of mast cells was significantly higher in the lateral portion of the TH than the medial portion of the TH. The numbers of mast cells increased on days 7-10 after the start of the experiment, and significantly decreased on days 14-21. Histamine-positive neurons and fibers in the TH and IC also had the same changes. Bleeding of the IC occurred exclusively around arteries, and perivenous bleeding was absent. Albumin exudation and suppression of EBA expression of capillaries were found in the spongy lesions of the TH and IC. The role of histamine in selective vulnerability of the TH and IC in experimental thiamine-deficient encephalopathy was not supported. Findings in the present study suggest that the spongy change is a primary event, and vascular changes are secondary.

[Change in nucleic acid synthesis in the spleen of rats with acute deficit of vitamin B1 during immunogenesis]. / Izmenenie sinteza nukleinovykkh kislot v selezenke krys s ostrym defitsitom vitamina B1 pri immunogeneze.

After four administrations of human blood serum gamma-globulin (intraperitoneally, once during 4 days at a dose of 1 mg and the following reimmunization within 60 days at a dose of 3-4 mg) distinct incorporation of 14C-thymidine (specific activity 54 muCi/mumole, by 5 muCi per an animal, 2 hrs before decapitation, intraperitoneally) into rat spleen DNA was detected within 4 days of immunogenesis. Acute oxythiamine B1 avitaminosis (single subcutaneous administration of oxythiamine at a dose of 400 mg/kg) led to a decrease in the label incorporation into spleen DNA of immunized rats by 70% (p less than 0.01) as compared with control immunized animals, while incorporation of 14C-thymidine was unaltered in intact rats. Content of RNA was not distinctly altered in spleen of B1-deficient rats within the 4 days of immunogenesis. Acute deficiency of vitamin B1 thus impaired synthesis of DNA in spleen during development of immunogenesis.

[Effect of acute vitamin B1 deficiency caused by oxythiamine on protein biosynthesis in the spleen during immunogenesis]. / Vliianie ostrogo defitsita vitamina B1, vyzvannogo oksitiaminom, na biosintez belka v selezenke pri immunogeneze.

Distinct increase in incorporation of 14C-protein hydrolysate of chlorella (5 microCi per an animal of 150 g body mass) into proteins of rat spleen was observed within 4 days after a single or repeated immunization of rats (180-200 g of body mass) with human blood serum gamma-immunoglobulin at a dose of 30-40 mg per 1 kg of animal mass. Under conditions of acute B1-avitaminosis caused by a single administration of hydroxythiamine at a high dose of 400 mg/kg incorporation of the label into spleen proteins of the immunized rats was decreased within 72 hrs. Thus, acute hydroxythiamine-produced B1-avitaminosis inhibited induction of protein biosynthesis stimulated during immunogenesis.

[Immunological disorders in vitamin A and B1 deficiencies]. / Immunologicheskie narusheniia pri defitsite vitaminov A i B1.

Atrophy of the thymus, mitogen-induced inhibition of T-lymphocyte blast-transformation, and decrease of the humoral immune response were observed in rats with alimentary vitamin A deficiency. Vitamin B1 deficiency induced by administration of oxythiamine led to the thymus atrophy, inhibition of T-lymphocyte blastogenic response in mice, and to suppression of delayed hypersensitivity in guinea pigs.

[Alkaline phosphatase and transketolase activity during immunogenesis in the lymphoid organs of rats with vitamin B1 deficiency caused by oxythiamine]. / Aktivnsot' shchelochnoi fosfatazy i transketolazy v limfoidnykh organakh krys s nedostatochnost'iu vitamina B1, vyzvannoi oksitiaminom, pri immunogeneze.

A study was made of the effect of different doses of hydroxythiamine on activity of alkaline phosphatase and transketolase in rat spleen and thymus during primary and secondary administration of human gamma-globulin. Immunization of rats significantly raised alkaline phosphatase activity in both organs. Administration of a large single dose of hydroxythiamine (400 mg/kg bw) reduced transketolase activity in the thymus and spleen in both immunized and non-immunized animals and did not change alkaline phosphatase activity in these organs.

Role of the B vitamins in the immune response.

Studies conducted in our laboratory relating to the development of immune processes in B vitamin deficiency states of experimental animals have been reviewed. 1. The significant participation of certain of these nutritional factors in the production of circulating antibodies to a variety of antigens and the manifestation of delayed hypersensitivity reactions, including the rejection of tissue transplants, have been described. 2. Investigations on the mode of action of pyridoxine and pantothenic acid have demonstrated a marked reduction in the production of antibody-forming cells following antigenic stimulation in both deficiency states. The metabolism of antigen appeared to be normal. However, these two vitamins seem to function at different loci in the development of the immune process. Whereas pyridoxine appears to be necessary for the production of "C1" units from serine which are required for the biosynthesis of nucleic acids, it seems likely that pantothenic acid is involved in the secretion of newly-synthesized proteins into the extracellular compartment.

[Change in the activity of specific and nonspecific immunity mechanisms in vitamin B1 deficiency]. / Izmenenie aktivonsti nekotorykh mekhanizmov spetsificheskogo i nespetsificheskogo immuniteta pri B1-vitaminnoi nedostatochnosti.

The effect of thiamin deficiency on the immune response and activity of some mechanisms of natural immunity was studied in experimental mature rats. Thiamin deficiency was simulated by a single injection of hydroxythiamin (thiamin antagonists). Hydroxythiamin markedly decreased the complement activity, phagocytic activity of peripheral blood leucocytes, serum bactericidal activity as well as antibody production in response to sheep red blood cells. On the contrary, lysozyme activity increased. Vitamin B1 deficiency reduced 14C-leucine incorporation activity in the liver proteins.

The inhibition of granuloma formation around Schistosoma mansoni eggs.

The effect of protein, calorie, and vitamin deficiency states on the granulomatous response of mice to eggs of S mansoni injected into the microvasculature of the lungs was studied. Moderate to severe protein and calorie deficiencies, as well as thiamine and vitamin C deficiencies, inhibited the host's granulomatous response to S mansoni eggs, which is a form of delayed-type hypersensitivity. The antivitamins neopyrithiamine hydrochloride and desoxypyridoxine hydrochloride were both highly effective in suppressing granuloma formation, but the former proved highly toxic at the doses utilized. Desoxypyridoxine hydrochloride given at a dose of .05 mg/mouse/day produced almost complete inhibition of granuloma formation; marked suppression was also present at fivefold dilution. This antivitamin appearts to be one of the most powerful immunosuppressive measures so far examined.