Diet-restricted Rats Research Articles

Intestinal morphology adjustments caused by dietary restriction improves the nutritional status during the aging process of rats

During the aging process, the body's systems change structurally and loss of function can occur. Ingesting a smaller amount of food has been considered a plausible proposal for increased longevity with the quality of life. However, the effects of dietary restriction (DR) during aging are still poorly understood, especially for organs of the digestive system. This study aimed to describe the body weight, oxidative status and possible morphological changes of the intestinal wall of rats submitted to DR during the aging process (7 to 18months old). Twelve 7-month-old male Wistar rats fed ad libitum since birth were assigned to two groups: control group (CG, n=6) fed ad libitum from 7 to 18months old; and dietary restriction group (DRG, n=6) fed 50% of the amount of chow consumed by the CG from 7 to 18months old. The body weight, feed and water intake were monitored throughout the experiment. Blood, periepididymal adipose tissue (PAT) and retroperitoneal adipose tissue (RAT), and the small intestine were collected at 18months old. The blood was collected to evaluate its components and oxidative status. Sections from the duodenum and ileum were stained with HE, PAS and AB pH2.5 for morphometric analyses of the intestinal wall components, and to count intraepithelial lymphocytes (IELs), goblet cells and cells in mitosis in the epithelium. DR rats showed a reduction in weight, naso-anal length, PAT, RAT and intestinal length; however, they consumed more water. Blood parameters indicate that the DR rats remained well nourished. In addition, they showed lower lipid peroxidation. Hypertrophy of the duodenal mucosa and atrophy of the ileal mucosa were observed. The number of goblet cells and IELs was reduced, but the mitotic index remained unaltered in both duodenum and ileum. In conclusion, 50% dietary restriction for rats from 7 to 18months old contributed to improving their nutritional parameters but, to achieve this, adjustments were required in the structure of the body weight and morphology of the small intestine.

Dietary restriction reduces age-related degeneration of stria vascularis in the inner ear of the rat

We report here beneficial effects of life-long dietary restriction on the progression of age-associated cochlear degeneration in female Sprague–Dawley rats. Thirty-month old rats on a 70% dietary restriction were compared to ad libitum fed age-matched rats, and three-month old adult rats. As expected, aged dietary restricted rats displayed about 20% higher survival rate than age-matched rats fed ad libitum. This difference was reflected also in the auditory system. In the dietary restricted group, 73% of the subjects had preserved auditory reflexes (Preyer), and only modest degeneration of the stria vascularis of the inner ear was observed. In contrast, aged ad libitum fed animals, of which only 15% had detectable Preyer reflexes, showed a marked thinning, cellular degeneration and loss of cell processes in the stria vascularis. The extent of loss of sensory hair cells (~24%) was similar in both the aged groups, and neither group showed a significant reduction in the number of spiral ganglion neurons across adult life-span. The observations thus demonstrate that dietary restriction delays age-related degradation of the auditory system. The results provide further insights into the mechanisms of strial presbycusis.

Neuroprotection provided by dietary restriction in rats is further enhanced by reducing glucocortocoids

Glucocorticoids (GC)—corticosterone (CORT) in rodents and cortisol in primates—are stress-induced hormones secreted by adrenal glands that interact with the hypothalamic pituitary axis. High levels of cortisol in humans are observed in neurodegenerative diseases, including Alzheimer's disease (AD) and Parkinson's disease (PD), as well as in diabetes, post-traumatic stress syndrome, and major depression. Experimental models of diabetes in rats and mice have demonstrated that reduction of CORT reduces learning and memory deficits and attenuates loss of neuronal viability and plasticity. In contrast to the negative associations of elevated GC levels, CORT is moderately elevated in dietary restriction (DR) paradigms which are associated with many healthy anti-aging effects including neuroprotection. We demonstrate here in rats that ablating CORT by adrenalectomy (ADX) with replenishment to relatively low levels (30% below that of controls) prior to the onset of a DR regimen (ADX-DR) followed by central administration of the neurotoxin, kainic acid (KA), significantly attenuates learning deficits in a 14-unit T-maze task. The performance of the ADX-DR KA group did not differ from a control group (CON) that did not receive KA and was fed ad libitum (AL). By contrast, the sham-operated DR (SHAM-DR KA) group, SHAM-AL KA group, and ADX-AL KA group demonstrated poorer learning behavior in this task compared to the CON group. Stereological analysis revealed equivalent DR-induced neuroprotection in the SH–DR KA and ADX-DR KA groups, as measured by cell loss in the CA2/CA3 region of the hippocampus, while substantial cell loss was observed in SH-AL and ADX-AL rats. A separate set of experiments was conducted with similar dietary and surgical treatment conditions but without KA administration to examine markers of neurotrophic activity, brain-derived neurotrophic factor (BDNF), transcriptions factors (pCREB), and chaperone proteins (HSP-70). Under these conditions, we noted elevations in both BDNF and pCREB in ADX DR rats compared to the other groups; whereas, HSP-70, was equivalently elevated in ADX-DR and SH–DR groups and was higher than observed in both SH-AL and ADX-AL groups. These results support findings that DR protects hippocampal neurons against KA-induced cellular insult. However, this neuroprotective effect was further enhanced in rats with a lower-than control level of CORT resulting from ADX and maintained by exogenous CORT supplementation. Our results then suggest that DR-induced physiological elevation of GC may have negative functional consequences to DR-induced beneficial effects. These negative effects, however, can be compensated by other DR-produced cellular and molecular protective mechanisms.

P.5.b.004 Cognition and hippocampal plasticity in 5XFAD mouse model of Alzheimer's disease

Dietary restriction (DR) is known to have potential health benefits including enhanced resistance of neurons to excitotoxic, oxidative and metabolic insults, cancer, stress, diabetes, reduced morbidity, and increased life span. In the present study, we examined the effect of DR (alternate day feeding regimen) on neurogenesis, expression of immature neuronal marker polysialic acid neural cell adhesion molecule (PSA-NCAM) and neurotrophic factors from different brain regions such as subventricular zone (SVZ), subgranular zone (SGZ) of hippocampus, median eminence arcuate (ME-ARC) region of hypothalamus, and piriform cortex (PIR) of adult male rats and further challenged ad libitum fed (AL) and DR rats with pilocarpine to induce excitotoxic injury. The quantitative analysis of bromodeoxyuridine (BrdU) labeling revealed a significant increase in the proliferation rate of neuronal progenitor cells from discrete brain regions in DR rats with and without pilocarpine induced seizures as compared to AL rats. DR significantly enhanced the expression of PSA-NCAM and neurotrophic factors, brain-derived neurotrophic factor (BDNF) and neurotrophin-3 (NT-3). There was a marked reduction in neuronal cell death in SVZ and PIR cortex after pilocarpine administration in DR rats. These results add to the accumulating evidence that DR may be an effective intervention to enhance the resistance of brain to excitotoxic injury.

STAT3/NF-κB interactions determine the level of haptoglobin expression in male rats exposed to dietary restriction and/or acute phase stimuli

Haptoglobin is a constitutively expressed protein which is predominantly synthesized in the liver. During the acute-phase (AP) response haptoglobin is upregulated along with other AP proteins. Its upregulation during the AP response is mediated by cis-trans interactions between the hormone-responsive element (HRE) residing in the haptoglobin gene and inducible transcription factors STAT3 and C/EBP β. In male rats that have been subjected to chronic 50% dietary restriction (DR), the basal haptoglobin serum level is decreased. The aim of this study was to characterize the trans-acting factor(s) responsible for the reduction of haptoglobin expression in male rats subjected to 50% DR for 6weeks. Protein-DNA interactions between C/EBP and STAT families of transcription factors and the HRE region of the haptoglobin gene were examined in livers of male rats subjected to DR, as well as during the AP response that was induced by turpentine administration. In DR rats, we observed associations between the HRE and C/EBPα/β, STAT5b and NF-κB p50, and the absence of interactions between STAT3 and NF-kB p65. Subsequent induction of the AP response in DR rats by turpentine administration elicited a normal, almost 2-fold increase in the serum haptoglobin level that was accompanied by HRE-binding of C/EBPβ, STAT3/5b and NF-kB p65/p50, and the establishment of interaction between STAT3 and NF-κB p65. These results suggest that STAT3 and NF-κB p65 crosstalk plays a central role while C/EBPβ acquires an accessory role in establishing the level of haptoglobin gene expression in male rats exposed to DR and AP stimuli.

An Investigation into the Effects of Aminoguanidine Treatment on the Plasma and Blood of Free-fed and Dietary-restricted Rats

An investigation has been made into the effect of oral aminoguanidine (50-60 mg kg-1 day-1) on the blood biochemistry of male Wistar rats which either had free access to food or were dietary-restricted (50% of the food consumed by the free access group). In control rats (i.e. without aminoguanidine treatment) three weeks of food restriction caused significant increases in plasma sodium and albumin and the erythrocyte count, haematocrit and haemoglobin. There were reductions in plasma calcium, phosphate, alkaline phosphatase activity, urea, triglycerides, creatinine, glucose and the red cell volume. Similar effects of food restriction were observed in aminoguanidine-treated rats. Aminoguanidine ingestion in free-fed animals caused a reduction in plasma creatinine concentration. In dietary-restricted rats, aminoguanidine ingestion reduced plasma sodium and total plasma proteins (largely as a result of a decline in albumin), and increased plasma urea concentrations. Aminoguanidine was added to plasma of control rats in-vitro to determine whether it interfered with the assay of urea and creatinine. At concentrations of 0.1 to 10 mg mL-1, aminoguanidine had no effect on urea determinations. However, aminoguanidine significantly reduced the apparent concentration of plasma creatinine by between 7 to 81%. The changes in plasma analytes in aminoguanidine-treated rats may be indicative of minor hepatic perturbations or kidney function, but the data also imply that prior nutritional state is a determinant of aminoguanidine effects.

The Effect of Aminoguanidine Treatment, With or Without Food Restriction on the Liver, Stomach and Small Intestine of the Rat

A comparative investigation has been made into the effects of aminoguanidine treatment (60 mg kg-1 day-1) with or without dietary restriction (i.e. 50% reduction in food intake) on the protein, RNA and DNA composition of the liver, small intestine and stomach of young rats (0.1 kg body weight). After 3 weeks of dietary restriction the wet weights of the liver and small intestine decreased by 56 and 52%, respectively. There were significant reductions (approx 50%) in total hepatic and intestinal protein, RNA and DNA. Changes in ratios of RNA/protein, RNA/DNA and protein/DNA were only significant for intestinal RNA/DNA, where a 15% reduction was observed. In contrast, stomach wet weight and total protein content were unaltered by dietary deprivation. Stomach RNA and DNA contents were reduced by only 18-21%, and the protein/DNA ratio increased by 22%. Similar responses of liver, small intestine and stomach to dietary deprivation were observed in aminoguanidine-treated rats. Aminoguanidine-treatment of rats on an unrestricted diet for three weeks had no effect on the wet weights, total protein, RNA or DNA contents of the liver, stomach or small intestine. In dietary-restricted rats, and stomach were unaffected by the treatment. However, aminoguanidine treatment of dietary-restricted rats caused significant increases in the amounts of intestinal protein, RNA and DNA by approximately 15%. The treatment abolished the dietary restriction-induced decrease in total intestinal DNA/body weight. The wet weights of the lung, diaphragm, kidney, spleen and testes of both fed and dietary-restricted rats were also unaffected by aminoguanidine.(ABSTRACT TRUNCATED AT 250 WORDS)

Peroxisome Proliferator-Activated Receptor-Gamma Expression in the Lung Tissue of Obese Rats

PurposeObesity is a risk factor for asthma and type II diabetes. Peroxisome proliferator-activated receptor (PPAR)-γ has been suggested to regulate inflammatory responses in diabetes and asthma. We investigated whether PPAR-α, PPAR-γ, adiponectin receptors (AdipoR1, AdipoR2), leptin, and tumor necrosis factor (TNF)-α are expressed in rat lung tissues and whether the expression differs between obese Otsuka Long-Evans Tokushima Fatty (OLETF) and lean Long Evans Tokushima Otsuka (LETO) rats.Materials and MethodsObese and lean rats were given with a high fat diet or a 30% restricted diet for 32 weeks, and their blood glucose levels and weights were monitored. After 32 weeks, mRNA levels of PPAR-α, PPAR-γ, AdipoR1, AdipoR2, leptin, and TNF-α in lung tissues were measured using real time PCR.ResultsPPAR-α, PPAR-γ, AdipoR1, AdipoR2, leptin, and TNF-α were expressed in both obese and lean rat lung tissues. Increased serum glucose levels on intraperitoneal glucose tolerance testing and a higher weight gain at 32 weeks were observed in OLETF control rats compared to OLETF diet restricted rats. PPAR-γ expression was markedly elevated in obese control and diet restricted rats compared to lean rats, although PPAR-γ expression in obese rats was not affected by diet restriction. Leptin was highly expressed in OLETF rats compared to LETO rats. TNF-α expression was enhanced in OLETF control rats compared LETO diet restricted rats, and decreased by diet restriction. PPAR-α, AdipoR1, and AdipoR2 expression were not significantly different between obese and lean rats.ConclusionPPAR-γ was highly expressed in the lung tissues of obese rats and may be a novel treatment target for regulating lung inflammation associated with obesity.

Muscle Wasting in Aged, Sarcopenic Rats Is Associated with Enhanced Activity of the Ubiquitin Proteasome Pathway

Among the hallmarks of aged organisms are an accumulation of misfolded proteins and a reduction in skeletal muscle mass ("sarcopenia"). We have examined the effects of aging and dietary restriction (which retards many age-related changes) on components of the ubiquitin proteasome system (UPS) in muscle. The hindlimb muscles of aged (30 months old) rats showed a marked loss of muscle mass and contained 2-3-fold higher levels of 26S proteasomes than those of adult (4 months old) controls. 26S proteasomes purified from muscles of aged and adult rats showed a similar capacity to degrade peptides, proteins, and an ubiquitylated substrate, but differed in levels of proteasome-associated proteins (e.g. the ubiquitin ligase E6AP and deubiquitylating enzyme USP14). Also, the activities of many other deubiquitylating enzymes were greatly enhanced in the aged muscles. Nevertheless, their content of polyubiquitylated proteins was higher than in adult animals. The aged muscles contained higher levels of the ubiquitin ligase CHIP, involved in eliminating misfolded proteins, and MuRF1, which ubiquitylates myofibrillar proteins. These muscles differed from ones rapidly atrophying due to disease, fasting, or disuse in that Atrogin-1/MAFbx expression was low and not inducible by glucocorticoids. Thus, the muscles of aged rats showed many adaptations indicating enhanced proteolysis by the UPS, which may enhance their capacity to eliminate misfolded proteins and seems to contribute to the sarcopenia. Accordingly, dietary restriction decreased or prevented the aging-associated increases in proteasomes and other UPS components and reduced muscle wasting.

Feature on Erythropoiesis in Dietary Restricted Rats

We attempted to characterize the influence of undernutrition on erythropoiesis in toxicity studies. Male rats were divided into the following 5 groups: dietary restriction groups in which feeding was restricted by 33% or 66% for 14 days (R33 and R66); phlebotomy groups in which 1% or 4% of total blood volume was removed by serial phlebotomy for 14 days (PB01 and PB04); and a nontreated group (NT). Toxicological parameters such as hematology and blood chemistry were evaluated. The body weight gains in the R33 and phlebotomy groups (PB01 and PB04) were similar and were less than that observed in the NT group. Decreases in peripheral blood reticulocytes, bone marrow erythroids and the unsaturated iron binding capacity (UIBC) were observed as changes that suppressed erythropoiesis in the R33 and R66 groups. However, increases in reticulocytes and UIBC were observed as opposite changes in the phlebotomy groups. In addition, an increase in the blood urea nitrogen level and a decrease in the serum alkaline phosphatase level were observed as changes reflecting poor nutrition in the phlebotomy groups. Decreased reticulocytes which are related to poor nutrition were not observed. However, increases in those cells as reflected by a loss of blood were observed in the phlebotomy groups. Even if undernutrition suppresses erythropoiesis, the ability of erythropoiesis to respond to a demand appears to be retained. In repeated dose toxicity studies, decreased food consumption is often observed in the drug administration groups. Our study results provide useful information for hematological evaluations in toxicity studies.

Dietary restriction suppresses inflammation and delays the onset of stroke in stroke-prone spontaneously hypertensive rats

Subjects with high blood levels of inflammatory markers and patients with chronic inflammatory disorders are at high risk for stroke. Dietary restriction (DR) suppresses systemic inflammation to deter age-related chronic diseases. To examine whether DR delays the onset of stroke, 10-week-old stroke-prone spontaneously hypertensive rats (SHRSP) were assigned to either a control ( ad libitum) or DR (50% diet of control) group, and day of stroke onset and lifespan were observed. DR markedly delayed the onset of stroke in SHRSP compared to control without affecting blood pressure. Day of stroke onset (median) in the control group was 34 days, whereas it was 70 days in the DR group. After 2 weeks of DR and before the onset of stroke, plasma levels of interleukin-1β (IL-1β), tumor necrosis factor-α (TNF-α), and monocyte chemoattractant protein-1 (MCP-1) and their mRNA expression levels in adipose tissue were significantly lower in the DR rats than in the control rats. Intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) mRNA expression levels in cerebrovascular endothelial cells (CVECs), and macrophage infiltration into brain were lower in the DR rats than in the control rats. IL-1β and TNF-α treatment in CVECs increased MCP-1, C-reactive protein, ICAM-1, and VCAM-1 mRNA and their protein levels in vitro. In conclusion, suppression of inflammation in response to DR may lead to a delay in the onset of stroke independent of any effect on blood pressure in SHRSP.

Altered thermoregulation via sensitization of A1 adenosine receptors in dietary-restricted rats

Evidence links longevity to dietary restriction (DR). A decrease in body temperature (T(b)) is thought to contribute to enhanced longevity because lower T(b) reduces oxidative metabolism and oxidative stress. It is as yet unclear how DR decreases T(b). Here, we test the hypothesis that prolonged DR decreases T(b) by sensitizing adenosine A(1) receptors (A(1)AR) and adenosine-induced cooling. Sprague-Dawley rats were dietary restricted using an every-other-day feeding protocol. Rats were fed every other day for 27 days and then administered the A(1)AR agonist, N(6)-cyclohexyladenosine (CHA; 0.5 mg/kg, i.p.). Respiratory rate (RR) and subcutaneous T(b) measured using IPTT-300 transponders were monitored every day and after drug administration. DR animals displayed lower RR on day 20 and lower T(b) on day 22 compared to animals fed ad libitum and displayed a larger response to CHA. In all cases, RR declined before T(b). Contrary to previous reports, a higher dose of CHA (5 mg/kg, i.p.) was lethal in both dietary groups. We next tested the hypothesis that sensitization to the effects of CHA was due to increased surface expression of A(1)AR within the hypothalamus. We report that the abundance of A(1)AR in the membrane fraction increases in hypothalamus, but not cortex of DR rats. These results suggest that every-other-day feeding lowers T(b) via sensitization of thermoregulatory effects of endogenous adenosine by increasing surface expression of A(1)AR. Evidence that diet can modulate purinergic signaling has implications for the treatment of stroke, brain injury, epilepsy, and aging.

Nutritional status alters saccharin intake and sweet receptor mRNA expression in rat taste buds

Sweet taste usually signifies the presence of caloric food. It is commonly accepted that a close association exists among sweet taste perception, preference, and nutritional status. However, the mechanisms involved remain unknown. To investigate whether nutritional status affects the preference for palatable solutions and alters sweet taste receptor gene expression in rats, we measured saccharin intake and preference using a two-bottle preference test, and changes in body weight, plasma leptin levels, and gene expression for the sweet taste receptor in taste buds in high-fat diet-induced obese rats and chronically diet-restricted rats. We found that the consumption and preference ratios for 0.01 and 0.04 M saccharin were significantly lower in the high-fat diet-induced obese rats than in the normal diet rats, while the serum leptin levels were markedly increased in obese rats. Consistent with the changes in saccharin intake, the gene expression level of the sweet taste receptor T1R3 was significantly decreased in the high-fat diet-induced obese rats compared with the control rats. By contrast, the chronically diet-restricted rats showed remarkably enhanced consumption and preference for 0.04 M saccharin. The serum leptin concentration was decreased, and the gene expression of the leptin receptor was markedly increased in the taste buds. In conclusion, our results suggest that nutritional status alters saccharin preference and the expression of T1R3 in taste buds. These processes may be involved in the mechanisms underlying the modulation of peripheral sweet taste sensitivity, in which leptin plays a role.

Long-term dietary restriction influences plasma ghrelin and GOAT mRNA level in rats

The objective of this study was to examine the effect of chronic dietary restriction on the physical characteristics of the intestine and gut-derived satiety hormone production. Male Wistar rats (8 weeks) were randomized to ad libitum (AL) or 35% dietary restriction (DR) for 5 months. At the end of the study, physical measurements were made on the intestine and satiety hormone secretion and mRNA expression determined. A comparison group of young, growing AL rats (5 weeks) was also examined. The adult DR rats gained less weight over 5 months and had lower fat mass than adult AL rats ( p < 0.05). The weight of the small intestine as a percentage of total body weight was greater in adult DR compared to adult AL but lower than young AL rats. Compared to AL, DR down-regulated proglucagon and cholecystokinin mRNA in the duodenum and ghrelin mRNA in the stomach of adult rats but was not different from young AL. Ghrelin-O-acyltransferase (GOAT) mRNA in the stomach was up-regulated 21-fold in adult AL rats compared to young AL and 14-fold compared to adult DR rats. Total and des-acyl ghrelin was approximately 50% higher in adult DR and young AL rats compared to adult AL. Plasma leptin and insulin were lower in adult DR and young AL rats compared to adult AL. Our findings suggest that long-term energy deficits continue to drive up ghrelin levels which may have profound implications for practical implementation of DR as an anti-aging or anti-obesity strategy in humans.

Interactive effect of excitotoxic injury and dietary restriction on neurogenesis and neurotrophic factors in adult male rat brain

Dietary restriction (DR) is known to have potential health benefits including enhanced resistance of neurons to excitotoxic, oxidative and metabolic insults, cancer, stress, diabetes, reduced morbidity, and increased life span. In the present study, we examined the effect of DR (alternate day feeding regimen) on neurogenesis, expression of immature neuronal marker polysialic acid neural cell adhesion molecule (PSA-NCAM) and neurotrophic factors from different brain regions such as subventricular zone (SVZ), subgranular zone (SGZ) of hippocampus, median eminence arcuate (ME-ARC) region of hypothalamus, and piriform cortex (PIR) of adult male rats and further challenged ad libitum fed (AL) and DR rats with pilocarpine to induce excitotoxic injury. The quantitative analysis of bromodeoxyuridine (BrdU) labeling revealed a significant increase in the proliferation rate of neuronal progenitor cells from discrete brain regions in DR rats with and without pilocarpine induced seizures as compared to AL rats. DR significantly enhanced the expression of PSA-NCAM and neurotrophic factors, brain-derived neurotrophic factor (BDNF) and neurotrophin-3 (NT-3). There was a marked reduction in neuronal cell death in SVZ and PIR cortex after pilocarpine administration in DR rats. These results add to the accumulating evidence that DR may be an effective intervention to enhance the resistance of brain to excitotoxic injury.

A novel dietary restriction method for group-housed rats: weight gain and clinical chemistry characterization

Laboratory rodents are usually fed ad libitum. Moderate dietary restriction decreases mortality and morbidity compared with ad libitum feeding. There are, however, problems in achieving dietary restriction. Traditional methods of restricted feeding may interfere with the diurnal rhythms of the animals and are not compatible with group-housing of rodents. We have invented a novel method, the diet board, for restricting the feed intake of laboratory rats. The use of the diet board moderately decreased weight gain of rats when compared with ad libitum-fed animals. The diet board retarded skeletal growth only minimally, whereas major differences were found in body fat depositions. Serum free fatty acid, triglyceride and cholesterol values were lower in diet-restricted rats, while the opposite was true for serum creatine kinase. There were no differences in total protein, albumin or alanine aminotransferase. Moreover, differences in interindividual variances in parameters were not detected between the groups; hence this study could not combine the diet board with reduction potential. The diet board provides mild to moderate dietary restriction for group-housed rats and is unlikely to interfere with the diurnal eating rhythm. The diet board can also be seen as a cage furniture item, dividing the open cage space and increasing the structural complexity of the environment. In conclusion, the diet board appears to possess refinement potential when compared with traditional methods of dietary restriction.

Differences between Molecular Mechanisms Involved in the Regulation of Haptoglobin Gene Expression during the Acute Phase Response and Dietary Restriction

Haptoglobin is a glycoprotein involved in the acute phase response. Previously we reported that haptoglobin gene expression was up-regulated during dietary restriction in young female rats. The present study aimed at determining whether chronic dietary restriction affects haptoglobin blood levels through changing levels and/or activities of IL-6-related transcription factors STAT and C/EBP in the liver as is the case during the acute phase response. To this end, we compared a female Wistar rat model of 50% 6-week-long dietary restriction with the standard laboratory model for the acute phase response induced by turpentine administration. During the turpentine-induced acute phase response, the transitory 5.4-fold increase of rat haptoglobin expression was accompanied by a prominent rise of serum IL-6 concentration and the increased binding of STAT3 and 35kD C/EBPβ/LAP transcription factors to the haptoglobin gene hormone-responsive element. Results obtained after immunoblotting and DNA affinity chromatography (using hormone-responsive element) suggest that the stable 1.7-fold increase of serum haptoglobin level during dietary restriction was the result of increased amounts and activities of constitutive transcription factors C/EBPα and STAT5b, and to a smaller extent of STAT3. When dietary restriction rats were administered turpentine, a 8.7-fold increase in haptoglobin expression was followed by a considerable increase in the amount and hormone-responsive element binding activity of STAT3 but not 35kD C/EBPβ/LAP. We concluded that haptoglobin gene up-regulation during chronic dietary restriction was regulated by different mechanisms than during the acute phase response, and that it depended on the amount(s) and activit(ies) of transcription factor(s) that characterize low-grade inflammatory conditions.

Importance of Starting Age for Myelotoxicity Study in Dietary Restricted Rats

The aim of this study was to prove our hypothesis that adult rats with lowering of body weight gain, rats at 12 weeks of age as an example, are suitable for evaluation of myelotoxicity. Age-related differences between young rats (6-week-old study) and adult rats (12-week-old study) were analyzed in hematological examination values. The data of the young rats were reprinted from our previous report (Miyata et al., 2009) since our hypothesis was verified by comparison with that previous report. Several experimental groups were defined for the 12-week-old study as well as for the 6-week-old study; these included 5-fluorouracil (5-FU) treated groups receiving 12, 15 and 18 mg/kg/day (FU12, FU15 and FU18), pair-feeding groups (R12, R15 and R18 receiving the same amount of food as in the FU12, FU15 and FU18 groups, respectively) and a nontreated control group. Numerous hematologic and bone marrow parameters in the 5-FU treated groups were comparable to those in the corresponding pair-feeding groups in both age studies. Generally, the influences of undernutrition were more apparent in the young rats than in the adult rats. Histopathological examinations showed a decrease in hematopoiesis in the bone marrow in the 5-FU treated and pair-feeding groups. No apparent differences were observed in the decreased hematopoiesis between the 5-FU treated and pair-feeding groups in the 6-week-old study, but a difference between these groups was noted in the 12-week-old study; decreased hematopoiesis was more frequently noted in the 5-FU treated groups. These facts suggest that adult rats are more suitable than young rats for evaluation of 5-FU-induced myelotoxicity.

Evaluation of Myelotoxicity in Dietary Restricted Rats

The purpose of this study was to clarify the effect of decreased food consumption on evaluation of myelotoxicity in routine general toxicity studies. Male rats were divided into the following 7 groups: 12, 15, and 18 mg/kg 5-fluorouracil (5-FU) treatment groups (FU12, FU15 and FU18); dietary restriction groups (R12, R15 and R18 receiving the same amount of food as the rats in the FU12, FU15 and FU18 groups, respectively); and a nontreated control group (NT). We compared the changes in body weight, hematology and the results of cytological analyses of bone marrow and histopathology among the groups after administration and recovery periods of 14 and 7 days, respectively. At the end of the administration period, the FU15 and FU18 groups showed decreases in many hematologic and bone marrow parameters that were all similar to those in the corresponding dietary restriction groups (R15 and R18). A granulocyte abnormality (polyploidy: frequency of 1% or less) was also observed in all 5-FU treated groups. At the end of the recovery period, increases in the reticulocyte and platelet counts and extramedullary hematopoiesis of the spleen were observed in the 5-FU treated groups. These results indicate that the results of general toxicity studies in rats should be evaluated in consideration of dietary restriction effects when food consumption is decreased at about 30-40% or more. Careful morphological observation of hemocytes would be helpful in distinguishing the effect of a drug from that of dietary restriction in relation to hematological and bone marrow parameters. Performance of a recovery test to determine the reactive response of hematopoiesis is also recommended.

Influence of Zinc Deficiency to the Rats Infected with Babesia rodhaini

Zinc deficiency induces a wide range of disorders including immunodeficiency. It is known that microbial infections occur with a high frequency in the zinc-deficient hosts, but the study on the correlation between parasitic infection and zinc status in hosts is scarcely performed. We observed that the influence of zinc deficiency to the rats infected with Babesia rodhaini. Experiments of B. rodhaini infection were conducted using zinc-deficient (ZD; eat ad libitum or 10 g/day on the ZD diet), zinc-adequate (ZA; eat ad libitum on the ZA diet), and diet-restricted (DR; eat 7 g/day on the ZA diet) rats. The findings in this study suggested that the zinc deficiency had deleterious effects on the hemodynamics and mortality of the rats infected with B. rodhaini.