Jejunal Crypt Cells Research Articles

Efficacy of soy protein concentrate replacing animal protein supplements in mucosa-associated microbiota, intestinal health, and growth performance of nursery pigs.

This study investigated the effects of using soy protein concentrate (SPC) to replace animal protein supplements on mucosa-associated microbiota, intestinal health, and growth performance of nursery pigs. Fifty-six newly weaned pigs (BW= 6.4±0.6kg) were allotted to 5 treatments in a randomized complete block design. Pigs were fed for 35d in 3 phases (P; 1, 2, 3) for 10, 12, 13d, respectively. Dietary treatments were: (1) basal diet with fish meal (P1: 4%, P2: 2%, and P3: 1%), poultry meal (P1: 10%, P2: 8%, and P3: 4%), and blood plasma (P1: 4%, P2: 2%, and P3: 1%), where SPC replacing none (NC); (2) basal diet with SPC replacing fish meal (RFM); (3) basal diet with SPC replacing poultry meal (RPM); (4) basal diet with SPC replacing blood plasma (RBP); and (5) basal diet with SPC replacing all animal protein supplements (PC). Growth performance was recorded for each phase. Pigs were euthanized on d 35 to collect jejunal mucosa and tissue to evaluate intestinal health and microbiota, and ileal digesta to measure apparent ileal digestibility (AID) of nutrients. Data were analyzed using the MIXED procedure of SAS. Overall, RFM, RPM, and RBP did not affect growth performance, whereas PC decreased (P<0.05) ADG and ADFI. The RPM increased (P<0.05) Prevotella stercorea and decreased (P<0.05) Helicobacter rappini. The PC decreased (P<0.05) H.rappini, whilst increasing (P<0.05) Prevotella copri, Propionibacterium acnes, and Pelomonas aquatica. The RFM tended to increase (P=0.096) immunoglobulin A in the jejunum. The PC tended to decrease (P=0.078) jejunal crypt cell proliferation. There were no differences in the villus height, AID of nutrients, intestinal inflammation, and intestinal oxidative stress among treatments. In conclusion, SPC can replace fish meal, poultry meal, or blood plasma individually without affecting growth performance and intestinal health, and AID of nutrients of nursery pigs. Particularly SPC replacing poultry meal benefitted intestinal health by reducing H.rappini and increasing P.stercorea. However, SPC replacing all three animal protein supplements reduced growth of nursery pigs mainly by reducing feed intake.

Heat stress disrupts intestinal stem cell migration and differentiation along the crypt–villus axis through FAK signaling

During heat stress (HS), the intestinal epithelium suffers damage due to imbalance of tissue homeostasis. However, the specific mechanism by which intestinal stem cells (ISCs) migrate and differentiate along the crypt–villus axis to heal lesions upon insult is unclear. In our study, C57BL/6 mice and IPEC-J2 cells were subjected to normal ambient conditions (25 °C for 7 days in vivo and 37 °C for 18 h in vitro) or 41 °C. The results showed that HS impaired intestinal morphology and barrier function. The numbers of ISCs (SOX9+ cells), mitotic cells (PCNA+ cells), and differentiated cells (Paneth cells marked by lysozyme, absorptive cells marked by Villin, goblet cells marked by Mucin2, enteroendocrine cells marked by Chromogranin A, and tuft cells marked by DCAMKL1) were reduced under high temperature. Importantly, BrdU incorporation confirmed the decreased migration ability of jejunal epithelial cells exposed to 41 °C. Furthermore, intestinal organoids (IOs) expanded from jejunal crypt cells in the HS group exhibited greater growth disadvantages. Mechanistically, the occurrence of these phenotypes was accompanied by FAK/paxillin/F-actin signaling disruption in the jejunum. The fact that the FAK agonist ZINC40099027 reversed the HS-triggered inhibition of IPEC-J2 cell differentiation and migration further confirmed the dominant role of FAK in response to high-temperature conditions. Overall, the present investigation is the first to reveal a major role of FAK/paxillin/F-actin signaling in HS-induced ISC migration and differentiation along the crypt–villus axis, which indicates a new therapeutic target for intestinal epithelial regeneration after heat injuries.

Early weaning leads to the remodeling of lipid profile in piglet jejunal crypt cells during post-weaning days.

Reportedly, proteins involved in lipid metabolism change significantly in the jejunal crypt cells of early-weaned piglets, but the exact lipid profile change remains uncertain. In the present study, 32 piglets weaned at 21 d of age were randomly divided into 4 groups with 8 replicates. The jejunal crypt cells of a group of piglets on the post-weaning day (PWD) 1, 3, 7, and 14 were isolated per time point. Crypt cell lipid profiles were analyzed using ultra-high-performance liquid chromatography coupled with hybrid quadrupole time-of-flight mass spectrometry. This study showed that piglets suffered the greatest weaning stress on PWD 3 in terms of the lowest relative weight of the small intestine, the highest relative weight of the spleen, and the highest levels of malondialdehyde, cholesterol, and low-density lipoprotein cholesterol. The lipid profile of jejunal crypt cells including carnitine, sulfatide, sphingomyelin, hexosylceramide, and ceramide greatly changed after weaning, especially between PWD 3 and 14 (P < 0.05). The differential lipid species between these 2 d were mainly involved in the glycerophospholipid metabolism pathway. In addition, potential lipid biomarkers for weaning stress in crypt cells such as phosphatidylcholine (PC) (9:0/26:1), PC (17:0/18:2), carnitine (24:0), carnitine (22:0), sphingomyelin (d14:1/22:0), PC (P-18:0/18:4), phosphatidylethanolamine (P-16:0/20:4), phosphatidylinositol (15:1/24:4), and dihexosylceramide (d14:1/26:1) were identified. The changes in lipid profile might be related to the inflammation caused by early weaning. These findings might provide new therapeutical targets for intestinal dysfunctions caused by weaning stress.

Pharmacologic Ascorbate Enhances the Therapeutic Index of ATM-Inhibitor Based Chemoradiation for Colorectal Cancer

Ataxia telangiectasia mutated protein (ATM) is one of the key sensors of DNA damage and specific inhibitors of ATM are potent radiosensitizers. However, their clinical utility with radiation (RT) is limited because they lack tissue specificity and increase normal tissue injury. Pharmacologic (high dose) ascorbate (P-AscH-) selectively increases oxidative stress in tumors while functioning as a donor antioxidant and reducing RT damage in normal tissues. We hypothesized that P-AscH- could enhance the therapeutic index of ATM-inhibitor based chemoradiation (CRT) for colorectal cancer (CRC) by simultaneously enhancing efficacy and reducing RT bowel injury.Human HCT116, SW480, and HT29 and murine CT26 and MC38 CRC models were used. Clonogenic survival was assessed following single-fraction RT (2-8 Gy) +/- P-AscH- (5 pM/cell) +/- veliparib (PARP), VE821 (ATR), or KU60019 (ATM). Catalase expression was induced using HCT116 cells expressing a doxycycline inducible catalase transgene. DNA double strand breaks (DSBs) were quantified using neutral comet assays 0-24 hours post RT. Cell cycle phases were assessed using flow cytometry. ATM and pATM localization were assessed using IF. Jejunal toxicity was assessed using IHC in fixed tissues following single fraction (10 Gy) whole abdominal RT in c57bl/6 mice. Tumor growth delay was assessed following RT (5 Gy x 3) +/- drug treatment in unilateral flank tumors.Veliparib, VE821, and KU60019 were potent radiosensitizers in HCT116, SW480, HT29, MC38, and CT26 CRC tumor models and P-AscH- further reduced clonogenic survival with DRIs in all lines except for HT29. In contrast, P-AscH- enhanced survival of cultured HUVEC and FHs-74 cells exposed to RT. Enhanced cell kill with P-AscH- is H202 mediated as it is completely attenuated by inducible catalase expression. P-AscH- significantly increased the number of DNA DSBs in tumors after RT in vitro. Despite the increase in DNA DSBs, P-AscH-significantly decreased nuclear localization of activated pATM after RT and significantly decreased the fraction of cells in G2/M phases of the cell cycle. In vivo, RT + P-AscH- + KU60019 induced more tumor growth delay/clearance than all other combinations in unilateral MC38 or HCT116 flank tumors. Finally, P-AscH- significantly reduced loss of jejunal crypt cell density, epithelial architecture, and markers of lipid and protein oxidation following whole abdominal RT.P-AscH- selectively enhances the efficacy of ATM-based CRT in CRC tumor models while simultaneously decreasing RT-mediated small bowel toxicity. In tumors, P-AscH- enhances DNA DSBs by stimulating an H202 flux and prevents activation of DNA repair pathways and cell cycle checkpoints by inhibiting RT-induced activation of ATM. Selective radioprotectors like P-AscH- could facilitate the clinical translation ATM inhibitors as radiosensitizers.

516 Late-Breaking: Iturin a Protects the Intestinal Mucosal Integrity Against E. coli Containing the Gene Coding for Heat-stable Enterotoxin B

Abstract Enterotoxigenic E. coli causes severe infectious diarrhea with high morbidity and mortality in weanling pigs mainly through the production of heat-stable enterotoxins. Iturin A, originally isolated from the insect Hyalophora cecropia, exerts strong antibiotic activity against Gram-positive and Gram-negative bacteria. Thus, our study investigated whether iturin A could protect the intestinal epithelium from the insults of E. coli Rosetta containing the gene coding for heat-stable enterotoxin b (STb-Rosetta). Twenty-eight piglets were divided into four groups and were orally administered PBS (control), 0.16 g/kg BW iturin A, 2 × 109 CFU STb-Rosetta, and 0.16 g/kg BW iturin A + 2 × 109 CFU STb-Rosetta respectively for 8 days. The intestinal epithelial morphology and barrier function were assessed by H&amp;E staining and Ussing chamber, and the growth advantage in enteroids derived from jejunal crypts of piglets in each group was used to evaluate the intestinal stem cell (ISC) activity. The results showed that iturin A increased the average daily gain (ADG) and average daily feed intake (ADFI) of piglets exposed to STb-Rosetta (P &amp;lt; 0.05). Meanwhile, STb-Rosetta-induced decrease in jejunum weight and damage to jejunal morphology and barrier function were significantly improved after iturin A supplementation (P &amp;lt; 0.05). Furthermore, the jejunal crypt cells from piglets in the iturin A + STb-Rosetta group had greater growth advantages of ISCs compared with the STb-Rosetta group, including enteroid forming efficiency, surface area, budding efficiency, and branching coefficient (P &amp;lt; 0.05). Our results indicate that iturin A protects against STb-Rosetta-induced intestinal mucosal injury, which provides an intervention strategy that regulates the function of ISCs in the high turnover-rate crypt-villous axis under ETEC infection. This study was supported by the National Natural Science Foundation of China (31872389, 32072777) and Basic and Applied Basic Research Foundation of Guangdong Province (2019B1515210021).

Pharmacologic ascorbate enhances the therapeutic index of ATM-inhibitor based chemoradiation for colorectal cancer.

3609 Background: Ataxia telangectasia mutated protein (ATM) is one of the key sensors of DNA damage and specific inhibitors of ATM are potent radiosensitizers. However, their clinical utility with radiation (RT) is limited because they lack tissue specificity and increase normal tissue injury. Pharmacologic (high dose) ascorbate (P-AscH-) selectively increases oxidative stress in tumors while functioning as a donor antioxidant and reducing RT damage in normal tissues. We hypothesized that P-AscH- could enhance the therapeutic index of ATM-inhibitor based chemoradiation (CRT) for colorectal cancer (CRC) by simultaneously enhancing efficacy and reducing RT bowel injury. Methods: Human HCT116, SW480, and HT29 and murine CT26 and MC38 CRC models were used. Clonogenic survival was assessed following single-fraction RT (2-8 Gy) +/- P-AscH- (5 pM/cell) +/- veliparib (PARP), VE821 (ATR), or KU60019 (ATM). Catalase expression was induced using HCT116 cells expressing a doxycycline inducible catalase transgene. DNA double strand breaks (DSBs) were quantified using neutral comet assays 0-24 hours post RT. Cell cycle phases were assessed using flow cytometry. ATM and pATM localization were assessed using IF. Jejunal toxicity was assessed using IHC in fixed tissues following single fraction (10 Gy) whole abdominal RT in c57blj/6 mice. Tumor growth delay was assessed following RT (5 Gy x 3) +/- drug treatment in unilateral flank tumors. Results: Veliparib, VE821, and KU60019 were potent radiosensitizers in HCT116, SW480, HT29, MC38, and CT26 CRC tumor models and P-AscH- further reduced clonogenic survival with DRIs in all lines except for HT29. In contrast, P-AscH- enhanced survival of cultured HUVEC and FHs-74 cells exposed to RT. Enhanced cell kill with P-AscH- is H202 mediated as it is completely attenuated by inducible catalase expression. P-AscH- significantly increased the number of DNA DSBs in tumors after RT in vitro. Despite the increase in DNA DSBs, P-AscH-significantly decreased nuclear localization of activated P-ATM after RT and significantly decreased the fraction of cells in G2/M phases of the cell cycle. In vivo, RT + P-AscH- + KU60019 induced more tumor growth delay/clearance than all other combinations in unilateral MC38 or HCT116 flank tumors. Finally, P-AscH- significantly reduced loss of jejunal crypt cell density, epithelial architecture, and markers of lipid and protein oxidation following whole abdominal RT. Conclusions: P-AscH- selectively enhances the efficacy of ATM-based CRT in CRC tumor models while simultaneously decreasing RT-mediated small bowel toxicity. In tumors, P-AscH- enhances DNA DSBs by stimulating an H202 flux and prevents activation of DNA repair pathways and cell cycle checkpoints by inhibiting RT-induced activation of ATM. Selective radioprotectors like P-AscH- could facilitate the clinical translation ATM inhibitors as radiosensitizers.

Protective effects of a cystine and theanine mixture against acute radiation injury in rats

This study aims to examine the effects of cystine and theanine (CT), which increases glutathione biosynthesis, on the survival rate and acute radiation injury of the small intestine and bone marrow using a rat model. CT pre-treatment (280 mg/kg for 5 days) significantly improved weight loss and survival rate of rats as compared with the control group after 5 Gy. CT pre-treatment significantly increased the rate of mucosa and crypt length, and decreased the number of apoptotic cells, TUNEL and cleaved caspase-3 positive cells, while increasing the number of mitotic cells and Ki-67 positive cells in jejunal crypts and villi compared to control rats post-irradiation. CT also suppressed bone marrow cell loss and reduced the number of apoptotic cells in bone marrow. These results suggest a protective effect of CT pre-treatment for acute injury after irradiation through apoptosis inhibition and increased proliferative activity in jejunal crypt cells and bone marrow cells.

Hydrolyzed wheat gluten alleviates deoxynivalenol-induced intestinal injury by promoting intestinal stem cell proliferation and differentiation via upregulation of Wnt/β-catenin signaling in mice

Disintegration of the intestine caused by deoxynivalenol (DON), which is a fungal metabolite found in cereal grain-based human and animal diets, triggers severe intestinal inflammatory disease. Hydrolyzed wheat gluten (HWG) can promote the development of intestine. Therefore, HWG was administered orally to male mice on 1–14 days, and DON was administered to them on 4–11 days. Feed, water intake and body weight were recorded all over the experimental period. Blood samples were collected then the mice were sacrificed to collect the jejunum for crypt isolation and culture. The intestinal morphology was observed by electron microscopy, and Western blotting was used to investigate intestinal stem cell (ISC) proliferation and differentiation, as well as the primary regulatory mechanism of the Wnt/β-catenin signaling. The results showed that HWG increased the average daily gain and average daily water intake of mice under DON-induced injury conditions, and increased the jejunum weight, villous height in the jejunum, and promoted jejunal crypt cell expansion. The DON-induced decrease in Wnt/β-catenin activity, the expression of Ki67, PCNA and KRT20 were rescued by HWG in the jejunum, crypt and enteroid, as well as the number of goblet cells and Paneth cells. Furthermore, HWG increased jejunum diamine oxidase (DAO) activity. In conclusion, HWG alleviates DON-induced intestinal injury by enhancing ISC proliferation and differentiation in a Wnt/β-catenin-dependent manner.

Effects of chito-oligosaccharide supplementation with low or medium molecular weight and high degree of deacetylation on growth performance, nutrient digestibility and small intestinal morphology in weaned pigs

Chito-oligosaccharide (COS) supplementation in nursery diets has been found to improve piglets’ performance. However, little is known on which COS characteristics are responsible for its effect. The present study was designed to further study how COS characteristics: molecular weight (MW) and degree of deacetylation (DD); affect the growth performance, nutrient digestibility, and small intestinal modifications in weaning pigs. For this purpose, 48 weanling pigs were divided into 4 groups (12 animals per group) and received either a basal diet or a basal diet supplemented with 150 mg/kg of COS which differed in MW and DD: 8 kDa and ~90% DD (8 kDa~90%), 65 kDa and ~80% DD (65 kDa~80%), and 65 kDa and ~90% DD (65 kDa~90%). The experiment lasted 56 days. There were treatment effects on pigs’ body weight gain (BWG), average daily gain (ADG), average daily feed intake (ADFI), blood urea nitrogen (BUN), digestibility of crude protein, fat and calcium, small intestinal morphology and crypt cell proliferation (P < .05, respectively). Throughout the study, pigs supplemented with 8 kDa~90% COS showed significantly enhanced BWG, ADG, and ADFI when compared to those supplemented with 65 kDa~80% COS (P < .05, respectively). The enhanced growth performances in pigs supplemented with 8 kDa~90% COS was supported by an increase in ADFI (P < .05), and improved ileal digestibility of crude protein and crude fat (P < .01, respectively). Pigs supplemented with 8 kDa~90% COS also had increased absorption capacity as indicated by elongated villus height and increased villus height to crypt depth ratio of the small intestinal segments (P < .05, respectively), and more active cell division as indicated by increased Ki-67 marker at jejunal crypt cells (P < .05) on day 28 and 56 of the experiment. These data suggest that 8 kDa~90% might be the most effective form of COS to improve feed intake, and promote nutrient digestibility and absorption when supplemented at 150 mg/kg in the diet.

Effects of dietary levels of chito-oligosaccharide on ileal digestibility of nutrients, small intestinal morphology and crypt cell proliferation in weaned pigs

There is continued need for novel agents to improve intestinal function in weaned pigs. Some evidence suggests that chito-oligosaccharide (COS) supplements may enhance pigs’ intestinal function after weaning. The present study was designed to examine the effect of COS supplementation on growth performance, nutrient digestibility and small intestinal functions in weaned pigs as an effective alternative to antibiotic addition in post-weaning diets. For the experiment, weanling pigs were divided into 5 groups (13–14 animals per group) and received either a basal diet, a supplemented diet with 75, 150 or 225mg/kg COS, or a supplemented diet with 110mg/kg lincomycin for 56 days. Growth, feed efficiency, hematological and biochemical profiles, nutrient's ileal digestibility, small intestinal morphology and crypt cell proliferation were measured at 28 and 56 days of the experiment. Pigs supplemented with 150mg/kg COS or lincomycin showed: (i) consistently more digestible ileal contents (e.g. crude protein, crude fat, ash, calcium, and phosphorus), (ii) increased absorption capacity (e.g. increased villus height and the villus height/crypt depth ratio for three intestinal segments) on day 28 of the experiment and (iii) more active cell division (as indicated by Ki-67 marker of duodenal and jejunal crypt cells) on day 56 of the experiment (P<0.05, respectively). These data suggest that 150mg/kg COS might be a useful dietary supplement to promote nutrient absorption and digestibility efficiency.

Effects of Golden Apple Snail (Pomacea Canaliculata, Lamarck) Shell Particle Size on Growth Performance, Carcass Quality, Bone Strength and Small Intestinal Histology in Thai Native Chickens (Pradu Hang Dum Chiangmai 1)

&lt;p class="1Body"&gt;To study whether the particle size of golden apple snail (&lt;em&gt;Pomacea Canaliculata&lt;/em&gt;, Lamarck) shell induces negative effects on the growth performance, carcass quality, bone strength and small intestinal histology in Thai native chickens (Pradu Hang Dum Chiangmai 1), 192 chickens, both male and female, were divided into 4 groups with 4 replicates of 12 chickens each at 5 weeks of age. The control group received limestone as a source of calcium. The experimental groups received diets containing golden apple snail shell particles, with sizes ranging from 0:50 to 1:00, from 1.00 to 1.70, and from 1.70 to 2.80 mm as a calcium source. Feed intake and weight were measured weekly, and carcass quality, tibial bone strength and small intestinal histology were observed at 16 weeks of age. The data of growth performance, carcass quality, tibial bone strength and small intestinal histology did not present negative results after feeding any of the golden apple snail shell sizes. On the contrary, a slightly higher weight gain was observed in the group consuming the 1.00 to 1.70 mm snail shell particles during the period from the 13th to the 16th week. Improved carcass quality and pectoralis major and tibial bone strength, as well as significantly increased duodenal villus surface and jejunal crypt cell numbers (p &amp;lt; 0.05) were observed in the group fed the 1.00 to 1.70 mm snail shell particles. These results suggest that golden apple snail shell particles between 1.00 and 1.70 mm can improve growth performance due to hypertrophied intestinal function.&lt;/p&gt;

Tu1271 Differential Mechanisms of EGF and GLP-2 Mediated Intestinal Epithelial Cell Proliferation

Introduction: Both GLP-2 and EGF can induce intestinal epithelial cell (IEC) growth. We previously showed that both growth factors (GFs) decrease with total parenteral nutrition (TPN) administration, and both exogenous GFs can prevent TPN-associated atrophy. However, the precise mechanisms driving this protective effect remain incompletely understood. This study investigated the role of PI3K/AKT and Wnt/β-catenin signaling mediating EC responses with TPN. Methods: C57BL/6 mice received enteral nutrition (Control) or TPN for 7 days. Wildtype, IEC-ErbB1KO and IEC-pik3r1KO (class IA subunit of PI3K) mice were treated with exogenous EGF or GLP-2. IEC proliferation was measured by PCNA staining. Nuclear and cytoplasm fractions were isolated from jejunal crypt cells and target proteins were measured by western blot. Results: TPN resulted in loss of both GFs expression and decreased EC proliferation. Exogenous EGF or GLP-2 restored IEC proliferation. We then explored differential mechanisms for EGF/GLP-2 responses. ErbB1: Both GFs actions weremarkedly decreased in IEC-ErbB1KOTPNmice, as well as in gefitinib (ErbB1 antagonist) treated TPN mice, demonstrating ErbB1-dependency for these GFs responses. P-AKT: Both GFs restored cytoplasmic p-Akt expression, but only EGF induced sustained nuclear pAKT expression. p-AKT expression was blocked in IEC-pi3kr1KO mice. However, in IECpi3kr1KO mice, GLP-2 induced EC proliferation but EGF. This indicated that GLP-2 acted by a PI3K-pAKT independent mechanism.Wnt-β-catenin: Increased cytoplasmic p-β-catenin (ser33/37) expression was found in TPNmice and both GFs decreased β-catenin ubiquination by preventing this increase. Nuclear p-β-catenin (ser552) expression was down-regulated in TPNmice and both GFs prevented this decline. Cytoplasmic p-GSK3βwas down-regulated in TPN mice and GLP-2 treatment prevented this decline but EGF did not. However, in IEC-pi3kr1KO TPN mice, nuclear p-β-catenin (ser552) and total cytoplasm β-catenin expression were maintained by GLP-2, whereas both β-catenin levels were still diminished in the presence of EGF. Stem cell: A marked loss of LGR5+ crypt cells (1% vs 20% , TPN vs Control) were observed in Lgr5-EGFP knock-in mice with TPN administration. EGF failed to prevent this loss, but LGR5+ was partially protected with GLP-2 (10%). Interestingly, although EC proliferation decreased in GLP-2 treated mice given gefinitib, LGR5+ crypts (10%) were still observed. Conclusions: Reduced EGF and GLP-2 signaling contributed to TPN-associated loss of EC proliferation. Both GFs required an intact ErbB1 signaling for their responses. However, EGF was dependent on PI3K/P-AKT signaling to promote EC proliferation, whereas GLP-2 was partially independent of the PI3K/p-AKT pathway to modulate EC proliferation and appeared to mediate proliferation viaWnt-β-catenin signaling.

Radiation protection from whole-body gamma irradiation (6.7 Gy): behavioural effects and brain protein-level changes by an aminothiol compound GL2011 in the Wistar rat

GL2011 is a naturally occurring thiol compound and a series of thiol compounds have been proposed as radioprotectors. Radioprotective efficacy of a triple intraperitoneal dose of GL2011 of 100 mg/kg body weight of Wistar rats, 30 min prior to and 3 and 6 h following irradiation (6.7 Gy) was evaluated. Four groups of animals were used, vehicle-treated non-irradiated (VN), GL2011-treated and irradiated (GI), GL2011-treated and non-irradiated (GN) and vehicle-treated and irradiated (VI) (n = 30 per group). The radioprotective efficacy of GL2011 was determined by measuring 28-day survival and intestinal crypt cell survival. Neuroprotection in terms of behaviour was evaluated using the behavioural observational battery, open field test and elevated plus maze paradigm. An RNA microarray was carried out in order to show differences at the RNA level between VI and VN groups. Brain protein changes were identified using a gel-based proteomics method and major brain receptor complex levels were determined by blue-native gels followed by immunoblotting. 28-Day survival rate in VI was 30 %, in GI survival was 93 %, survival of VN and GN was 100 %. Jejunal crypt cell survival was significantly enhanced in GI. Protein-level changes of peroxiredoxin-5, Mn-superoxide dismutase 2, voltage-dependent anion-selective channel protein 1, septin 5 and dopamine D2 receptor complex levels were paralleling radiation damage and protection. Taken together, the findings demonstrate that GL2011 improves survival rates and jejunal crypt survival, provides partial neuroprotection at the behavioural level and modulates proteins known to be involved in protection against oxidative stress-mediated cell damage.

Geraniin down regulates gamma radiation-induced apoptosis by suppressing DNA damage

Gamma ray irradiation triggers DNA damage and apoptosis of proliferating stem cells and peripheral immune cells, resulting in the destruction of intestinal crypts and lymphoid system. Geraniin is a natural compound extracts from an aquatic plant Nymphaea tetragona and possesses good antioxidant property. In this study, we demonstrate that geraniin rescues radiosensitive splenocytes and jejunal crypt cells from radiation-induced DNA damage and apoptosis. Isolated splenocytes from C57BL/6 mice treated with geraniin were protected against radiation injury of 2 Gy irradiation through the enhancement of the proliferation and attenuation of DNA damage. Also, geraniin inhibited apoptosis in radiosensitive splenocytes by reducing the expression level and immunoreactivity of proapoptotic p53 and Bax and increasing those of anti-apoptotic Bcl-2. In mice exposed to radiation, geraniin treatment protected splenocytes and intestinal crypt cells from radiation-induced cell death. Our results suggest that geraniin presents radioprotective effects by regulating DNA damage on splenocytes, exerting immunostimulatory capacities and inhibiting apoptosis of radiosensitive immune cells and jejunal crypt cells. Therefore, geraniin can be a radioprotective agent against γ-irradiation exposure.

Impacts of maternal selenium supply and nutritional plane on visceral tissues and intestinal biology in 180-day-old offspring in sheep1

Objectives were to investigate the effects of maternal Se supply and nutritional plane during gestation on offspring visceral tissues and indices of intestinal growth, vascularity, and function at 180 d of age. Rambouillet ewe lambs (n = 82, approximately 240 d of age; 52 ± 0.8 kg BW at breeding) were allocated to a 2 × 3 factorial arrangement of treatments. Treatments included dietary Se [adequate Se (ASe, 9.5 µg/kg BW) or high Se (HSe, 81.8 µg/kg BW)] initiated at breeding and nutritional plane [60% (restricted, RES), 100% (control, CON), and 140% (high, HI) of requirements] initiated at d 50 of gestation. Ewes were fed pelleted diets and housed individually indoors. At parturition, lambs were immediately removed and fed artificial colostrum for the first 20 h followed by ad libitum access to milk replacer. At 180 ± 2 d of age, lambs were euthanized and tissues were harvested. Birth weight was affected by nutritional treatments (P < 0.001), with decreased birth weight in RES and HI compared with CON. Offspring from RES and HI ewes had decreased (P = 0.07) blood volume compared with CON, and those born to HSe ewes had increased (P < 0.04) total visceral adiposity. Within offspring from CON ewes, those from HSe ewes had greater (P < 0.02) intestinal mass compared with ASe ewes. Within offspring from HSe ewes, both RES and HI had reduced (P ≤ 0.05) intestinal mass compared with CON. Jejunal capillary area density was greater (P = 0.08) in offspring from ewes fed HSe compared with ASe. In addition, area per capillary was greater (P ≤ 0.09) in CON compared with RES. Maternal nutritional plane tended (P ≤ 0.11) to alter total small intestinal vascularity, with lambs from CON being greater than RES. Expression of most mRNA for measured angiogenic factors and receptors was not altered (P ≤ 0.13) by maternal treatments; however, expression of glucagon-like peptide-2 (GLP-2) was decreased (P = 0.07) in offspring from RES compared with CON ewes. Offspring from ewes fed HI diets had increased (P = 0.08) jejunal mucosal maltase activity. In conclusion, maternal Se supply and nutritional plane during gestation resulted in measurable changes in offspring visceral tissues and intestinal biology, including perirenal fat, blood volume, intestinal mass, total jejunal crypt cell proliferation, area per capillary in jejunal villi, GLP-2 mRNA expression, and maltase activity at 180 d. Additional work is needed to determine impacts on intestinal function and nutrient uptake.

민들레 추출물의 방사선 방호효과

민들레는 한방에서 산화방지, 위장 점막 보호 및 담즙 등에 탁월한 효과가 있다고 알려져 있다. 민들레 잎과 뿌리에서 추출한 물질을 SD Rat에 투여한 후 10Gy 방사선 조사 후에 생존율, 혈액변화, 소장조직변화 및 간 조직 변화를 관찰하였다. 관찰결과 민들레 추출물을 투여한 실험군에서 생존율이 4배 이상 높았으며 백혈구, 혈소판 등의 혈액변화에서도 뚜렷한 차이가 보였다. 또한 소장 조직 관찰결과 소장점막세포 소멸 억제 및 소장 점막근판 조직의 염증과 공포가 감소되었다. 따라서 민들레 추출물이 방사선 조사에 따른 혈액구성세포와 소장 조직 손상에 대한 방사선 방호효과를 확인하였다. Dandelion(Taraxacum officinale), an oriental herbal medicine, has been shown to favorably affect choleretic, antioxidative and protection of gastric mucosa. The protective effects of common dandelion leaf and root extract were investigated by irradiation in Sprague-Dawley rats. Male SD rats, 6weeks, were orally injected with dandelion extract(100mg/kg) for 10days. Immediately after final injection, rats were whole body irradiated with 10Gy used irradiation facility(Elekta Linac, Sweden). At 24h-15days after irradiation, complete blood counted test and apoptosis in jejunal crypt cell. Stimulated recovery by the extract was observed in platelet but was not showed in the erythrocyte and leucocyte. The jejunal crypt cells were protected significantly(p<0.05) and the radiation-induced apoptosis was reduced(p<0.05). The survival rate were carried for 15days, the survival ratio was 15% and 85% for the control and experimental group. Observations intestinal inhibition of cell death, intestinal mucosa and tissue decreased systemic inflammation and vacuole. Base on these data, we propose that dandelion may be a useful radioprotector, especially since it is a relatively nontoxic natural product.

Phloroglucinol Protects Small Intestines of Mice from Ionizing Radiation by Regulating Apoptosis-Related Molecules

Phloroglucinol (PG) is a phenolic compound isolated from Ecklonia cava, a brown algae abundant on Jeju island, Korea. Previous reports have suggested that PG exerts antioxidative and cytoprotective effects against oxidative stress. In this study, we confirmed that PG protected against small intestinal damage caused by ionizing radiation, and we investigated its protective mechanism in detail. Regeneration of intestinal crypts in the PG-treated irradiated group was significantly promoted compared with that in irradiated controls. The expression level of proapoptotic molecules such as p53, Bax, and Bak in the small intestine was downregulated and that of antiapoptotic molecules such as Bcl-2 and Bcl-X(S/L) was augmented in the PG-treated group. On histological observation of the small intestine, PG inhibited the immunoreactivity of p53, Bax, and Bak and increased that of Bcl-2 and Bcl-X(S/L). These results demonstrate the protective mechanisms of PG in mice against intestinal damage from ionizing radiation, providing the benefit of raising the apoptosis threshold of jejunal crypt cells.

In vitro and in vivo study of a nanoliposomal cisplatin as a radiosensitizer

Objective:To investigate the in vitro and in vivo radiosensitization effect of an institutionally designed nanoliposome encapsulated cisplatin (NLE-CDDP).Materials and methods:NLE-CDDP was developed by our institute. In vitro radiosensitization of NLE-CDDP was evaluated by colony forming assay in A549 cells. In vivo radiosensitization was studied with tumor growth delay (TGD) in Lewis lung carcinoma. The radiosensitization for normal tissue was investigated by jejunal crypt survival. The radiosensitization studies were carried out with a 72 h interval between drug administration and irradiation. The mice were treated with 6 mg/kg of NLE-CDDP or CDDP followed by single doses of 2 Gy, 6 Gy, 16 Gy, and 28 Gy. Sensitization enhancement ratio (SER) was calculated by D0s of cell survival curves for A549 cells, doses needed to yield TGD of 20 days in Lewis lung carcinoma, or D0s of survival curves in crypt cells in radiation alone and radiation plus drug groups.Results:Our NLE-CDDP could inhibit A549 cells in vitro with half maximal inhibitory concentration of 1.12 μg/mL, and its toxicity was 2.35 times that observed in CDDP. For in vitro studies of A549 cells, SERs of NLE-CDDP and CDDP were 1.40 and 1.14, respectively, when combined with irradiation. For in vivo studies of Lewis lung carcinoma, the strongest radiosensitization was found in the 72 h interval between NLE-CDDP and irradiation. When given 72 h prior to irradiation, NLE-CDDP yielded higher radiosensitization than CDDP (SER of 4.92 vs 3.21) and slightly increased injury in jejunal crypt cells (SER of 1.15 vs 1.19). Therefore, NLE-CDDP resulted in a higher TGF than did CDDP (4.28 vs 2.70) when SERs were compared between experiments in vivo and in jejunal crypt cell studies.Conclusions:Our NLE-CDDP was demonstrated to have radiosensitization with TGF of 4.28 when administrated 72 h prior to irradiation.

Norepinephrine Enhances Radiosensitivity in Rat Ileal Epithelial Cells

We previously reported that the apoptosis index in jejunal crypt cells after X irradiation was greater in spontaneously hypertensive rats than in Wistar-Kyoto rats. Moreover, these same cells showed a suppression of apoptosis when reserpine was administered to induce sympathetic dysfunction in spontaneously hypertensive rats or Wistar-Kyoto rats. Whether the hyperfunction of the sympathetic nervous system is involved in the high susceptibility of the jejunal crypt cells to radiation-induced apoptosis was the subject of this study. The effect of norepinephrine (NE) on cell survival was examined using the colony formation assay after X-ray irradiation of rat ileal epithelial cells (IEC-18). The addition of 1 µM NE decreased the surviving fraction of cells irradiated with 6 Gy from 37% to 8%. The radiosensitivity of IEC-18 cells was enhanced by the addition of 1 µM of NE. The irradiation and treatment with NE also resulted in an increased cellular apoptotic rate. These results showing enhanced radiosensitivity of rat ileal epithelial cells by NE suggest that NE may be one of the factors which aggravate acute radiation injury in the intestine.

Basic Fibroblast Growth Factor Suppresses Radiation-Induced Apoptosis and TP53 Pathway in Rat Small Intestine

The effect of basic fibroblast growth factor (bFGF) was studied in radiation-induced apoptosis in rat jejunal crypt cells. Six-week-old male Wistar rats were administered 4 mg/kg bFGF intraperitoneally 25 h before receiving 8 Gy whole-body X rays. The jejunum was removed for analysis from time 0 to 120 h after irradiation. Villus length in control rats declined steadily until 72 h, while in bFGF-treated rats the villi were longer than in the controls until 48 h. Crypt lengths were similar to villi. bFGF treatment increased Ki-67-positive cells in the jejunal crypt at 0, 24 and 48 h. The treatment with bFGF reduced the number of apoptotic cells per jejunal crypt to 23% and 10% of the control values at 3 and 6 h, respectively, and increased numbers of mitotic cells significantly at 48 and 72 h. bFGF decreased the levels of TP53, CDKN1A, Puma and Cleaved caspase 3 at 3 h as detected by Western blot analyses. Our results suggest that bFGF protected against acute radiation-induced injury by suppressing the crypt apoptotic cells including the stem cells and promoted crypt cell proliferation. The inhibition of apoptosis thus might be related to suppression of the TP53 pathway.