4-week Recovery Period Research Articles

Excessive eccentric exercise-induced overtraining model leads to endoplasmic reticulum stress in mice skeletal muscles

AimsThe present study verified the responses of selected endoplasmic reticulum (ER) stress proteins (i.e., BiP, ATF-6, pIRE1, pPERK, and peIF2alpha) in mice skeletal muscles after three different running overtraining (OT) protocols with same external load (i.e., intensity vs. volume), but performed in downhill, uphill and without inclination. Materials and methodsThe rodents were randomly divided into control (CT; sedentary mice), overtrained by downhill running (OTR/down), overtrained by uphill running (OTR/up) and overtrained by running without inclination (OTR) groups. The incremental load test and exhaustive test were used as performance parameters. Forty hours after the exhaustive test performed at the end of the OT protocols (i.e., at the end of week 8) and after a 2-week total recovery period (i.e., at the end of week 10), the extensor digitorum longus (EDL) and soleus muscles were removed and used for immunoblotting. Key findingsFor both skeletal muscle types, the OTR/down protocol increased the pIRE-1, pPERK and peIF2alpha, which were not normalized after the total recovery period. At the end of week 8, the other two OT protocols up-regulated the BiP, pPERK and peIF2alpha levels only for the soleus muscle. These ER stress proteins were not normalized after the total recovery period for the OTR/up group. SignificanceThe above findings suggest that the OTR/down protocol-induced skeletal muscle ER stress may be linked to a pathological condition in EDL and soleus muscles.

Excessive eccentric exercise leads to transitory hypothalamic inflammation, which may contribute to the low body weight gain and food intake in overtrained mice

Low body weight gain and food intake are related to exhaustive training and overtraining; however, the molecular mechanisms responsible for these alterations remain unknown. The main aim of this study was to evaluate the effects of running overtraining (OT) protocols performed downhill, uphill and without inclination on the inflammatory pathway in the mouse hypothalamus. The rodents were randomized into the control (C), overtrained by downhill running (OTR/down), overtrained by uphill running (OTR/up) and overtrained by running without inclination (OTR) groups. The body weights and food intake were recorded daily. The incremental load, exhaustive, rotarod and grip force tests were used to measure performance. At 36h after the grip force test was performed at the end of OT protocols (i.e., week eight) and/or after a 2-week total recovery period (i.e., week 10), the hypothalamus and gastrocnemius were extracted for immunoblotting analysis. In addition, the serum was used to determine cytokine and leptin concentrations. From week 0 to week 8, the OTR/down group exhibited decreased body weight and food intake, and the OTR/up group increased their food intake. At week 10, the OTR/down group exhibited increased body weight, while the OTR group decreased their food intake. The OTR/down group exhibited increased IL-1beta, IL-6, TNF-alpha, pSAPK/JNK and SOCS3 levels at week eight. The OTR/down, OTR/up and OTR groups exhibited increased IL-10 levels at week 10. The OTR/up group displayed increased pJAK2 levels at week eight. While the OTR/down group exhibited increased IL-1beta levels, the OTR/down and OTR/up groups exhibited increased IL-6 and TNF-alpha levels, but decreased IL-10 levels in the gastrocnemius at week eight. The three OT protocols increased the IL-1beta and IL-6 levels, but only the OTR/down and OTR/up groups had increased TNF-alpha levels in serum at week eight. The serum leptin levels were lower for the OTR group compared with the CT group at week eight. In conclusion, the OTR/down protocol induced transitory hypothalamic inflammation with concomitant reductions in the body weight and food intake. After the 2-week total recovery period, the OTR/down group had reversed the hypothalamic inflammation, with the concomitant normalization of the body weight and food intake.

Acute and sub-chronic toxicity study of the ethanol extract from leaves of Aralia elata in rats

Ethnopharmacological relevanceAralia elata Seem. (A. elata) is a well-known medicinal plant which has been used as a tonic, anti-arthritic and anti-diabetic agent in traditional Chinese medicine. This investigation aimed to evaluate the potential toxicological properties of the ethanol extract from leaves of A. elata, namely ethanol leaves extract (ELE), in rats by acute and sub-chronic toxicity studies. Materials and methodsIn the acute toxicity study, rats were orally administrated with ELE at doses of 1.00, 2.15, 4.64, and 10.00g/kg to determine the oral medial lethal dose (LD50). Abnormal behavior, toxic symptom, and death were observed for 14 consecutive days. In the sub-chronic toxicity study, rats were orally administrated with ELE at doses of 0, 60, 180, and 540mg/kg for 12 weeks and followed-up a 4-week recovery period. At the end of the treatment and recovery periods, the rats were sacrificed for hematological, biochemical, and histopathology analyses. ResultsThe acute toxicity study showed that oral administration of ELE induced the incidence of adverse effects. The death rate also increased in a dose-dependent manner. The LD50 value was 3.16g/kg for female rats, and 5.84g/kg for male rats, respectively. The sub-chronic toxicity study showed that daily oral administration of ELE induced no significant difference in food consumption. However, the body weight of male rats in high dose group increased slowly compared with the control group during the recovery period. The hematological and biochemical analysis showed that compared with the control group, HGB and MCV levels were significantly increased in ELE treatment groups at the end of the treatment period, while TP and GLB levels were significantly decreased at the end of the recovery period. The absolute and relative weight of thymus, brain and adrenal gland showed a significant difference in low or high dose group at the end of the treatment period, although no histological changes were observed in various organs. ConclusionThe results of this study provided evidence that oral administration of ELE at dose of 540mg/kg is safe in rats and may not exert severe toxic effects.

The effects of visceral obesity and androgens on bone: trenbolone protects against loss of femoral bone mineral density and structural strength in viscerally obese and testosterone-deficient male rats.

In males, visceral obesity and androgen deficiency often present together and result in harmful effects on bone. Our findings show that both factors are independently associated with adverse effects on femoral bone structure and strength, and trenbolone protects rats from diet-induced visceral obesity and consequently normalises femoral bone structural strength. In light of the rapidly increasing incidence of obesity and osteoporosis globally, and recent conjecture regarding the effects of visceral adiposity and testosterone deficiency on bone health, we investigated the effects of increased visceral adipose tissue (VAT) mass on femoral bone mineral density (BMD), structure and strength in normal weight rats with testosterone deficiency. Male Wistar rats (n = 50) were fed either standard rat chow (CTRL, n = 10) or a high-fat/high-sugar diet (HF/HS, n = 40). Following 8 weeks of feeding, rats underwent sham surgery (CTRL, n = 10; HF/HS, n = 10) or orchiectomy (HF/HS + ORX, n = 30). Following a 4-week recovery period, mini-osmotic pumps containing either vehicle (CTRL, n = 10; HF/HS, n = 10; HF/HS + ORX, n = 10), 2.0 mg kg day(-1), testosterone (HF/HS + ORX + TEST, n = 10) or 2.0 mg kg day(-1) trenbolone (HF/HS + ORX + TREN, n = 10) were implanted for 8 weeks of treatment. Dual-energy X-ray absorptiometry and three-point bending tests were used to assess bone mass, structure and strength of femora. Diet-induced visceral obesity resulted in decreased bone mineral area (BMA) and content (BMC) and impaired femoral stiffness and strength. Orchiectomy further impaired BMA, BMC and BMD and reduced energy to failure in viscerally obese animals. Both TEST and TREN treatment restored BMA, BMC, BMD and energy to failure. Only TREN reduced visceral adiposity and improved femoral stiffness and strength. Findings support a role for both visceral adiposity and testosterone deficiency as independent risk factors for femoral osteoporosis, adverse bone geometry and impaired bone strength in male rats. Trenbolone may be a more effective candidate for androgen replacement therapy than testosterone in viscerally obese testosterone-deficient males.

REGENERATION IN MINT (MENTHA × PIPERITA) CRYOPRESERVED APICES: CAN THE CRYOPRESERVATION TECHNIQUE, REGENERATION MEDIUM COMPOSITION AND GENOTYPE AFFECT THE FINAL RESULT?

One of the most reliable methods for long-term conservation is cryopreservation, mainly due to its capability to guarantee the genetic stability of the preserved material. During the development of cryopreservation protocols the composition of the growth recovery medium is of vital importance in order to obtain fully viable structures without callus formation. Two cryopreservation techniques (droplet-vitrification and encapsulation-dehydration) and three different recovery media were compared with shoot apices of two mint genotypes (MEN 186 and MEN 198). The medium was supplemented with: 1) 0.5 mg/L 6-benzylaminopurine; 2) 0.5 mg/L 6-dimethylallylamino-purine + 0.1 mg/L α-naphthalene acetic acid; or 3) 0.5 mg/L 6-benzylaminopurine + 0.1 mg/L α-naphthalene acetic acid. Survival and regeneration (shoot growth) were studied after a 4-week recovery period. Although the recovery media did not influence survival in any of the two genotypes or protocols, the regeneration percentage observed was affected by the growth regulators. When the encapsulation-dehydration protocol was employed this influence was detected even in control apices (not subjected to cooling in liquid nitrogen), while with the droplet-vitrification method recovery medium had an effect only on cryopreserved apices. In all these cases, the medium without auxin (only with cytokinin) resulted in higher recovery rates, i.e., organized growth was favoured.

Lethal and sub-lethal chronic effects of the herbicide diuron on seagrass

Photosystem II herbicides from agricultural sources have been detected throughout nearshore tropical habitats including seagrass meadows. While PSII herbicides have been shown to inhibit growth in microalgae at low concentrations, the potential impacts of chronic low concentration exposures to seagrass health and growth have not been investigated. Here we exposed two tropical seagrass species Halodule uninervis and Zostera muelleri to elevated diuron concentrations (from 0.3 to 7.2μgl−1) over a 79-day period followed by a 2-week recovery period in uncontaminated seawater. PAM fluorometry demonstrated rapid effect of diuron on photosystem II (PSII) in both seagrass species at 0.3μgl−1. This effect included significant inhibition of photosynthetic efficiency (ΔF/Fm′) and inactivation of PSII (Fv/Fm) over the 11 week exposure period. Significant mortality and reductions in growth was only observed at the highest exposure concentration of 7.2μgl−1 diuron. However, biochemical indicators demonstrated that the health of seagrass after this prolonged exposure was significantly compromised at lower concentrations. For example, the drop in C:N ratios (0.6μgl−1) and reduced δ13C (1.7μgl−1) in seagrass leaves indicated reduced C-assimilation from photosynthesis. Critically, the energetic reserves of the plants (as measured by starch content in the root-rhizome complex) were approximately halved following diuron exposure at and above 1.7μgl−1. During the 2-week recovery period, the photosynthetic capacity of the seagrass improved with only plants from the highest diuron treatment still exhibiting chronic damage to PSII. This study shows that, although seagrass may survive prolonged herbicide exposures, concentrations ≥0.6μgl−1 diuron equivalents cause measureable impacts on energetic status that may leave the plants vulnerable to other simultaneous stressors. For example, tropical seagrasses have been heavily impacted by reduced light from coastal flood plumes and the effects on plant energetics from light limitation and diuron exposure (highest in flood plumes) are very similar, potentially leading to cumulative negative effects.

Biological responses in rats exposed to mainstream smoke from a heated cigarette compared to a conventional reference cigarette

The heated cigarette (HC) generates mainstream smoke by vaporizing the components of the tobacco rod using a carbon heat source at the cigarette tip. Mainstream smoke of HC contains markedly less chemical constituents compared to combusted cigarettes. Mainstream smoke from HC was generated under Health Canada Intense regimen and its biological effects were compared to those of Reference (3R4F) cigarettes, using nose-only 5-week and 13-week inhalation studies. In the 13-week study, SD rats were necropsied following exposure to mainstream smoke from each cigarette at 200, 600 or 1000 µg wet total particulate matter/L for 1 h/day, 7 days/week or following a 13-week recovery period. Histopathological changes in the respiratory tract were significantly lesser in HC groups; e.g. respiratory epithelial hyperplasia in the nasal cavity and accumulation of pigmented macrophages in alveoli. After a 13-week recovery, the lesions were completely or partially regressed, except for accumulation of pigmented macrophages in alveoli, in both HC and 3R4F groups. In the 5-week study, SD rats were necropsied following exposure to mainstream smoke of either cigarette at 600 or 1000 µg/L for 1 h, two times/day (with 30 min interval), 7 days/week or following a 4-week recovery period. Bronchoalveolar lavage fluid (BALF) analysis of neutrophil percentages and enzyme levels like γ-GT, ALP and LDH indicated that pulmonary inflammation was significantly less in HC groups compared to 3R4F groups. In conclusion, HC demonstrated significantly lower biological effects compared to 3R4F, based on the BALF parameters and histopathology.

Discordance in Recovery Between Altered Locomotion and Muscle Atrophy Induced by Simulated Microgravity in Rats

ABSTRACTExposure to a microgravity environment leads to adverse effects in motion and musculoskeletal properties. However, few studies have investigated the recovery of altered locomotion and muscle atrophy simultaneously. The authors investigated altered locomotion in rats submitted to simulated microgravity by hindlimb unloading for 2 weeks. Motion deficits were characterized by hyperextension of the knees and ankle joints and forward-shifted limb motion. Furthermore, these locomotor deficits did not revert to their original form after a 2-week recovery period, although muscle atrophy in the hindlimbs had recovered, implying discordance in recovery between altered locomotion and muscle atrophy, and that other factors such as neural drives might control behavioral adaptations to microgravity.

Safety evaluation of standardized allergen extract of Japanese cedar pollen for sublingual immunotherapy

Japanese cedar (JC) pollinosis is caused by Japanese cedar pollen (JCP) and most common seasonal allergic disease in Japan. Subcutaneous immunotherapy (SCIT) with allergen extract of JCP (JCP-allergen extract) is well established for JC pollinosis treatment with improvement of symptoms. However, major drawbacks for SCIT are repeated painful injections, frequent hospital visits and anaphylactic risk. Currently, sublingual immunotherapy (SLIT) has received much attention as an advanced alternative application with lower incidence of systemic reactions because the liquid or tablet form of allergen is placed under the tongue. The aim of this study was safety evaluation of standardized JCP-allergen extract currently developed for SLIT in JC pollinosis. JCP-allergen extract showed no potential genotoxicity. No systemic effects were observed in rats administered JCP-allergen extract orally for 26weeks followed by 4-week recovery period. Mild local reactions such as hyperplasia and increased globule leukocytes resulting from vehicle (glycerin)-induced irritation were observed in stomach. No-observed-adverse-effect level was greater than 10,000JAU/kg/day for systemic toxicity, equivalent to 300-fold the human dose. No local irritation was found in rabbits oral mucosae by 7-day sublingual administration. These results demonstrate the safe profile of standardized JCP-allergen extract, suggesting it is suitable for SLIT in JC pollinosis.

Cartilage oligomeric matrix protein and matrix metalloproteinase-3 expression in the serum and joint fluid of a reversible osteoarthritis rabbit model.

The main pathological characteristic of osteoarthritis (OA) is cartilage damage. We explored cartilage oligomeric matrix protein (COMP) and matrix metalloproteinase-3 (MMP-3) changes during articular cartilage injury and repair. Rabbits were randomly divided into the following: a blank control group; groups M1, M2, and M3, in which breaking was performed for 2, 4, and 6 weeks, respectively; and groups L1, L2, and L3, in which breaking was discontinued for 2, 4, and 6 weeks, respectively, following a 4-week recovery period. There are 7 rabbits in each group. The degree of cartilage damage in each group was scored (OA score). An enzyme-linked immunosorbent assay was used to detect COMP and MMP-3 levels in serum and joint fluid. The OA scores were 3.89 ± 2.31, 7.21 ± 2.31, and 10.88 ± 2.08 points in groups M1, M2, and M3, respectively (P < 0.05). COMP and MMP-3 levels were significantly higher in groups M1, M2, and M3 than in C. The OA score improved significantly following the 4-week recovery period (P < 0.05). COMP and MMP-3 levels began to decrease as the time following discontinuation of breaking increased, but were higher than in the control (P < 0.05). MMP- 3 and COMP levels were correlated with OA score (r > 0.7, P < 0.05). COMP and MMP-3 levels were correlated between joint fluid and serum (r = 0.899, r = 0.874, P < 0.05, respectively). Long-term joint breaking can cause articular cartilage damage. Doing some activites after the process can promote self-repair of articular cartilage. COMP and MMP-3 levels were associated with articular cartilage destruction and repair.

Effects of reduced food intake on the parameters of toxicity evaluation in dogs.

It is crucial to evaluate the variations in the toxicity parameters in experimental animals during the development of new drugs. Reduced food intake has been reported to have an impact on the toxicity parameters in rats; however, there are few reports of such studies in dogs. The aim of this study was to clarify the effects of reduced food intake on the general toxicity parameters and their reversibility in dogs. Male beagle dogs were fed 300 g/day of diet for 12 weeks in the control group, and 150 g/day for the first 8 weeks and 200 g/day for the subsequent 4 weeks in the low feeding group. During the following 4-week recovery period, the amount of feeding was set at 300 g/day. There were no clinical changes in any of the dogs. The low feeding group showed a body weight loss of 9.0%, 16.7% and 14.3% relative to the pre-test values at Week 4, 8 and 12, respectively. The following changes from the pre-test values and/or the control group in the examined parameters were observed in this group: decreased heart rate, prolonged PR interval on the ECG, decreased leukocyte count, and increased serum free fatty acid and γ-glutamyl transpeptidase levels. Significant changes of these parameters were not observed any more during the recovery period. This fact supports biological or physiological reaction to reduced food intake. These results are considered to represent useful information for toxicologists to distinguish between the direct effects of drugs and the changes attributable to reduced food intake.

Myopia Control during Orthokeratology Lens Wear in Children Using a Novel Study Design

To investigate the effect of overnight orthokeratology (OK) contact lens wear on axial length growth in East Asian children with progressive myopia. A prospective, randomized, contralateral-eye crossover study conducted over a 1-year period. We enrolled 26 myopic children (age range, 10.8-17.0 years) of East Asian ethnicity. Subjects were fitted with overnight OK in 1 eye, chosen at random, and conventional rigid gas-permeable (GP) lenses for daytime wear in the contralateral eye. Lenses were worn for 6 months. After a 2-week recovery period without lens wear, lens-eye combinations were reversed and lens wear was continued for a further 6 months, followed by another 2-week recovery period without lens wear. Axial eye length was monitored at baseline and every 3 months using an IOLMaster biometer. Corneal topography (Medmont E300) and objective refraction (Shin-Nippon NVision-K 5001 autorefractor) were also measured to confirm that OK lens wear was efficacious in correcting myopia. Axial length elongation and myopia progression with OK were compared with conventional daytime rigid contact lens wear. After 6 months of lens wear, axial length had increased by 0.04±0.06 mm (mean±standard deviation) in the GP eye (P=0.011) but showed no change (-0.02±0.05 mm) in the OK eye (P=0.888). During the second 6-month phase of lens wear, in the OK eye there was no change from baseline in axial length at 12 months (-0.04±0.08 mm; P=0.218). However, in the GP eye, the 12-month increase in axial length was significant (0.09±0.09 mm; P<0.001). The GP lens-wearing eye showed progressive axial length growth throughout the study. These results provide evidence that, at least in the initial months of lens wear, overnight OK inhibits axial eye growth and myopia progression compared with conventional GP lenses. Apparent shortening of axial length early in OK lens wear may reflect the contribution of OK-induced central corneal thinning, combined with choroidal thickening or recovery due to a reduction or neutralization of the myopiogenic stimulus to eye growth in these myopic children.

Acute and sub-chronic oral toxicity profiles of the aqueous extract of Cortex Dictamni in mice and rats

Ethnopharmacological relevanceCortex Dictamni is used in Chinese folk medicine for the treatment of jaundice, cough, rheumatism and some skin diseases; however, its possible toxicity has not been rigorously studied. The present investigation was carried out to evaluate the safety of Cortex Dictamni aqueous extract (CDAE) by acute and sub-chronic toxicity studies. Materials and methodsIn acute toxicity tests, seven groups of mice (n=5/group/sex) were orally treated with doses of 0, 28.7, 33.6, 39.7, 46.7, 54.9 and 64.6g/kg of CDAE and general behavior, adverse effects and mortality were recorded for up to 14 days. In sub-chronic toxicity assays, animals received CDAE by gavage at the doses of 0, 3.0, 6.0 or 12.0g/kg/day (n=10/group/sex) for 4 weeks and then followed for a 2-week recovery period. The biochemical, hematological and morphological parameters were determined. ResultsIn adult mice, single oral administrations of CDAE (0–64.6g/kg body weight) induced an increase in the incidence of general behavioral adverse effects. The mortality rate also increased with increasing dosage (LD50=48.2g/kg). In rats, daily single oral doses of CDAE were well tolerated behaviorally after 4 weeks and induced no significant changes in body weights. However, the absolute and relative liver weight at the end of both administration and recovery periods were significantly elevated, although the histological examination of various organs revealed no differences between the control and the treated groups. The hematological and biochemical parameters were significantly changed; lymphocytes, alanine transaminase, alkaline phosphatase levels showed a significant decrease while neutrophilic granulocyte, albumin, total cholesterol, glucose and blood urea nitrogen levels showed a significant increase, suggesting disturbances of hematopoiesis and liver and kidney functions. ConclusionsOverall, the acute toxicity of CDAE was not clearly observed. However, it is possible that CDAE has a selective toxicity considering the changes in some hematological and liver function parameters and the liver-body weight ratios in the sub-chronic oral toxicity study.

Bilateral contusion-compression model of incomplete traumatic cervical spinal cord injury.

Despite the increasing incidence and prevalence of cervical spinal cord injury (cSCI), we lack clinically relevant animal models that can be used to study the pathomechanisms of this injury and test new therapies. Here, we characterize a moderate cervical contusion-compression model in rats that is similar to incomplete traumatic cSCI in humans. We characterized the effects of 18-g clip-compression injury at cervical level C6 over an 8-week recovery period. Using Luxol fast blue/hematoxylin-eosin staining in combination with quantitative stereology, we determined that 18-g injury results in loss of gray matter (GM), white matter (WM), as well as in cavity formation. Magnetization transfer and T2-weighted magnetic resonance imaging were used to analyze lesion dynamics in vivo. This analysis demonstrated that both techniques are able to differentiate between the injury epicenter, subpial rim, and WM distal to the injury. Neurobehavioral assessment of locomotor function using Basso, Beattie, and Bresnahan (BBB) scoring and CatWalk revealed limited recovery from clip-compression injury at C6. Testing of forelimb function using grip strength demonstrated significant forelimb dysfunction, similar to the loss of upper-limb motor function observed in human cSCI. Sensory-evoked potentials recorded from the forelimb and Hoffman reflex recorded from the hindlimb confirmed the fore- and hindlimb deficits observed in our neurobehavioral analysis. Here, we have characterized a clip-compression model of incomplete cSCI that closely models this condition in humans. This work directly addresses the current lack of clinically relevant models of cSCI and will thus contribute to improved success in the translation of putative therapies into the clinic.

SNS01-T Modulation of eIF5A Inhibits B-cell Cancer Progression and Synergizes With Bortezomib and Lenalidomide

The high rates of recurrence and low median survival in many B-cell cancers highlight a need for new targeted therapeutic modalities. In dividing cells, eukaryotic translation initiation factor 5A (eIF5A) is hypusinated and involved in regulation of protein synthesis and proliferation, whereas the non-hypusinated form of eIF5A is a potent inducer of cell death in malignant cells. Here, we demonstrate the potential of modulating eIF5A expression as a novel approach to treating B-cell cancers. SNS01-T is a nonviral polyethylenimine-based nanoparticle, designed to induce apoptosis selectively in B-cell cancers by small interfering RNA–mediated suppression of hypusinated eIF5A and plasmid-based overexpression of a non-hypusinable eIF5A mutant. In this study, we show that SNS01-T is preferentially taken up by malignant B cells, inhibits tumor growth in multiple animal models of B-cell cancers without damaging normal tissues, and synergizes with the current therapies bortezomib and lenalidomide to inhibit tumor progression. The results collectively demonstrate the potential of SNS01-T as a novel therapeutic for treatment of a diverse range of B-cell malignancies.

Spinal growth modulation with posterior unilateral elastic tether in immature swine model

Background contextFusionless scoliosis surgery is frequently performed in children. Many studies have analyzed the effects of spinal growth modulation by tethering the anterior and anterolateral aspects of the spine in animal models. However, few studies have reported the disc health and spinal motion in spines with posterior unilateral elastic tethering. PurposeTo analyze the regional radiography, biochemistry, and histology of spinal motion segments fixed by posterior unilateral elastic tethering. Study designA randomized controlled trial. Outcome measuresPreoperative and postoperative radiographs of the spines were taken. After an 8-week recovery period, the spines were harvested en bloc and underwent radiographic, biochemical, and histologic analyses. MethodsFifteen 3-month-old swine were randomly divided into three groups. Instrumentation was performed posteriorly in the swine. In the elastic fixation (EF) group, five swine were instrumented on the left side of the lumbar vertebrae from L1 to L5 with pedicle screws that were connected with a unilateral elastic tether with tension to produce a curve on the spine. The same surgery was performed in the five animals of the metal rod fixation (MF) group, in which the screws were connected with metal rods and curves were established. In the control group, five animals were instrumented with five screws with no connecting cable. ResultsScoliosis and lordosis were created in the coronal and sagittal planes in both the EF and MF groups. On average, the Cobb angles were 12.16°±1.37° and 9.10°±2.02° (p=.023) in the coronal plane and 17.44°±11.29° and 5.32°±3.06° (p=.049) in the sagittal plane in the two groups, respectively. The vertebrae and discs wedged on the tethered side in the two groups showed no significant differences (p>.05). The thickness of end-plate epiphysis on the fixed side was significantly decreased in the two groups (p=.032 and p=.024). No apparent change was found in the gross morphology of the discs in the two groups. The distribution of collagen types I and II decreased and that of matrix metalloprotease-3 (MMP-3) increased in both the EF and MF groups. Additionally, the proteoglycan synthesis decreased in the two groups. ConclusionsUnilateral elastic tethering resulted in vertebral wedging and scoliosis. Although changes in collagen and MMP-3 distribution, proteoglycan synthesis, end-plate epiphysis, and disc thickness were observed, the tethered discs and end plates did not demonstrate gross morphologic signs of degeneration.

Mango and pomegranate polyphenolics in the modification of microbiota and short chain fatty acids in DSS‐induced colitis (1045.6)

Dysregulation of the interactions between gut microbiota and the immune system is speculated to be involved in triggering colitis. Gut microbiota can be modulated by dietary compounds such as fiber and polyphenolics. To evaluate the effects of mango and pomegranate polyphenolics on fecal microbiota and SCFAs production, rats were administered control, mango, or pomegranate juice, and were exposed to three cycles of 3% DSS followed by 2‐week recovery period. Fecal samples were collected to characterize the fecal microbiota composition using selected representative genus by qPCR and to measure SCFAs concentration. Mango and pomegranate juice decreased the level of pro‐inflammatory cytokines, such as IL‐1b and GM‐CSF in serum, and induced the level of IL‐10. Pomegranate juice induced compositional changes in the fecal microbiota. In particular, the anti‐inflammatory Ruminococcaceae significantly was increased by pomegranate juice in the feces but the concentrations of SCFAs were not significantly affected. In response to the mango juice, fecal SCFAs isovalerate and valerate were increased, while there was no significant change in the composition of the intestinal microbiota at the genus level. Polyphenolics decreased the TLR4 expression, but only mango increased the GPR43 expression. In summary, mango juice induced changes in the SCFAs production while pomegranate juice induced changes in the composition of microbiota.

Comparison of anti‐inflammatory mechanisms of mango (Mangifera indica L.) and pomegranate (Punica granatum L.) in DSS‐induced colitis in rats (372.8)

Ulcerative colitis, a chronic inflammation of the large intestine, may increase risk of human colorectal cancer. Several studies have focused on developing natural anti‐inflammatory products. Polyphenolics from mango (mainly gallotannins) and pomegranate (mainly ellagitannins) have been shown to have potent anti‐inflammatory properties. To determine the anti‐inflammatory effects and possible mechanisms of mango and pomegranate juice in DSS‐induced colitis, SD rats were administered control, mango, or pomegranate juice, and were exposed to three cycles of 3% DSS followed by 2‐week recovery period. The levels of protein involved in the mTOR pathway were analyzed by multiplex bead assay, while the transcriptional variation with the mTOR pathway were analyzed by low density PCR arrays. In addition, the levels of several miRNAs involved in the mTOR pathway were measured. Chronic colitis was caused by DSS in rats, and mango and pomegranate juice reduced colon inflammation compared to control juice. Mango juice suppressed the IGF‐1R‐AKT/mTOR signaling axis via up‐regulation of miR‐126 and down‐regulation of IR, while pomegranate decreased p70S6K by up‐regulating miR‐145 and down‐regulation of the MEK‐ERK1/2 pathway. These results suggest that polyphenolics of different predominant structure may differentially regulate inflammation‐involved pathways while attenuating DSS‐induced colitis.

Phenobarbital Induces Cell Cycle Transcriptional Responses in Mouse Liver Humanized for Constitutive Androstane and Pregnane X Receptors

The constitutive androstane receptor (CAR) and the pregnane X receptor (PXR) are closely related nuclear receptors involved in drug metabolism and play important roles in the mechanism of phenobarbital (PB)-induced rodent nongenotoxic hepatocarcinogenesis. Here, we have used a humanized CAR/PXR mouse model to examine potential species differences in receptor-dependent mechanisms underlying liver tissue molecular responses to PB. Early and late transcriptomic responses to sustained PB exposure were investigated in liver tissue from double knock-out CAR and PXR (CAR(KO)-PXR(KO)), double humanized CAR and PXR (CAR(h)-PXR(h)), and wild-type C57BL/6 mice. Wild-type and CAR(h)-PXR(h) mouse livers exhibited temporally and quantitatively similar transcriptional responses during 91 days of PB exposure including the sustained induction of the xenobiotic response gene Cyp2b10, the Wnt signaling inhibitor Wisp1, and noncoding RNA biomarkers from the Dlk1-Dio3 locus. Transient induction of DNA replication (Hells, Mcm6, and Esco2) and mitotic genes (Ccnb2, Cdc20, and Cdk1) and the proliferation-related nuclear antigen Mki67 were observed with peak expression occurring between 1 and 7 days PB exposure. All these transcriptional responses were absent in CAR(KO)-PXR(KO) mouse livers and largely reversible in wild-type and CAR(h)-PXR(h) mouse livers following 91 days of PB exposure and a subsequent 4-week recovery period. Furthermore, PB-mediated upregulation of the noncoding RNA Meg3, which has recently been associated with cellular pluripotency, exhibited a similar dose response and perivenous hepatocyte-specific localization in both wild-type and CAR(h)-PXR(h) mice. Thus, mouse livers coexpressing human CAR and PXR support both the xenobiotic metabolizing and the proliferative transcriptional responses following exposure to PB.

Studies on the toxicity and distribution of indium compounds according to particle size in sprague-dawley rats.

Objectives: The use of indium compounds, especially those of small size, for the production of semiconductors, liquid-crystal panels, etc., has increased recently. However, the role of particle size or the chemical composition of indium compounds in their toxicity and distribution in the body has not been sufficiently investigated. Therefore, the aim of this study was to examine the effects of particle size and the chemical composition of indium compounds on their toxicity and distribution. Methods: Male Sprague-Dawley rats were exposed to two different-sized indium oxides (average particle sizes under 4,000 nm [IO_4000] and 100 nm [IO_100]) and one nano-sized indium-tin oxide (ITO; average particle size less than 50 nm) by inhalation for 6 hr daily, 5 days per week, for 4 weeks at approximately 1 mg/m3 of indium by mass concentration. Results: We observed differences in lung weights and histopathological findings, differential cell counts, and cell damage indicators in the bronchoalveolar lavage fluid between the normal control group and IO- or ITO-exposed groups. However, only ITO affected respiratory functions in exposed rats. Overall, the toxicity of ITO was much higher than that of IOs; the toxicity of IO_4000 was higher than that of IO_100. A 4-week recovery period was not sufficient to alleviate the toxic effects of IO and ITO exposure. Inhaled indium was mainly deposited in the lungs. ITO in the lungs was removed more slowly than IOs; IO_4000 was removed faster than IO_100. IOs were not distributed to other organs (i.e., the brain, liver, and spleen), whereas ITO was. Concentrations of indium in the blood and organ tissues were higher at 4 weeks after exposure. Conclusions: The effect of particle size on the toxicity of indium compounds was not clear, whereas chemical composition clearly affected toxicity; ITO showed much higher toxicity than that of IO.