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Papers by JESSICA GUADALUPE CALVO DOMINGUEZ

Research paper thumbnail of TNF is a key mediator in sepsis‐induced intestinal barrier dysfunction but is independent of enterocyte NFκB

Research paper thumbnail of Epidermal growth factor treatment prevents intestinal injury in weanling mice with septic peritonitis

Research paper thumbnail of Plan de empresa Jeshia Construction

Según la demanda de proyectos solicitados contrataremos a más profesionales del sector de la cons... more Según la demanda de proyectos solicitados contrataremos a más profesionales del sector de la construcción.

Research paper thumbnail of Plan De Relaciones Publicas Para Agencia De Investigacion De Mercados

Research paper thumbnail of Intestine-Specific Deletion of Microsomal Triglyceride Transfer Protein Increases Mortality in Aged Mice

PLoS ONE, 2014

Background: Mice with conditional, intestine-specific deletion of microsomal triglyceride transfe... more Background: Mice with conditional, intestine-specific deletion of microsomal triglyceride transfer protein (Mttp-IKO) exhibit a complete block in chylomicron assembly together with lipid malabsorption. Young (8-10 week) Mttp-IKO mice have improved survival when subjected to a murine model of Pseudomonas aeruginosa-induced sepsis. However, 80% of deaths in sepsis occur in patients over age 65. The purpose of this study was to determine whether age impacts outcome in Mttp-IKO mice subjected to sepsis. Methods: Aged (20-24 months) Mttp-IKO mice and WT mice underwent intratracheal injection with P. aeruginosa. Mice were either sacrificed 24 hours post-operatively for mechanistic studies or followed seven days for survival. Results: In contrast to young septic Mttp-IKO mice, aged septic Mttp-IKO mice had a significantly higher mortality than aged septic WT mice (80% vs. 39%, p = 0.005). Aged septic Mttp-IKO mice exhibited increased gut epithelial apoptosis, increased jejunal Bax/Bcl-2 and Bax/Bcl-X L ratios yet simultaneously demonstrated increased crypt proliferation and villus length. Aged septic Mttp-IKO mice also manifested increased pulmonary myeloperoxidase levels, suggesting increased neutrophil infiltration, as well as decreased systemic TNFa compared to aged septic WT mice. Conclusions: Blocking intestinal chylomicron secretion alters mortality following sepsis in an age-dependent manner. Increases in gut apoptosis and pulmonary neutrophil infiltration, and decreased systemic TNFa represent potential mechanisms for why intestine-specific Mttp deletion is beneficial in young septic mice but harmful in aged mice as each of these parameters are altered differently in young and aged septic WT and Mttp-IKO mice.

Research paper thumbnail of Epidermal growth factor reduces the development of necrotizing enterocolitis in a neonatal rat model

American journal of physiology. Gastrointestinal and liver physiology, 2002

Necrotizing enterocolitis (NEC) is the most common gastrointestinal disease of prematurely born i... more Necrotizing enterocolitis (NEC) is the most common gastrointestinal disease of prematurely born infants. Maternal milk plays an important protective role against NEC development and is the major source of epidermal growth factor (EGF) for neonates. The aim of this study was to examine the effect of orally administered EGF on the incidence of NEC in a neonatal rat model. Newborn rats were artificially fed either with growth factor-free rat milk substitute (RMS) or RMS supplemented with 500 ng/ml of EGF (RMS+EGF). Experimental NEC was induced by exposure to asphyxia and cold stress. Development of NEC was evaluated by gross and histological scoring of damage in the ileum. Ileal EGF receptor (EGF-R), EGF, and transforming growth factor-alpha mRNA expression was assessed by RT competitive-PCR, and the EGF-R was localized by immunohistochemistry. EGF supplementation of formula reduced the incidence and severity of NEC in rats (13/16 RMS vs. 4/13 RMS+EGF). Ileal EGF-R mRNA expression was ...

Research paper thumbnail of Hepatic inflammatory mediators contribute to intestinal damage in necrotizing enterocolitis

American journal of physiology. Gastrointestinal and liver physiology, 2003

Necrotizing enterocolitis (NEC) is a common and devastating gastrointestinal disease of premature... more Necrotizing enterocolitis (NEC) is a common and devastating gastrointestinal disease of premature infants. Along with pathological effects in the ileum, severe NEC is often accompanied by multisystem organ failure, including liver failure. The aim of this study was to determine the changes in hepatic cytokines and inflammatory mediators in experimental NEC. The well-established neonatal rat model of NEC was used in this study, and changes in liver morphology, numbers of Kupffer cells (KC), gene expression, and histological localization of IL-18, TNF-alpha, and inducible nitric oxide synthase were evaluated. Intestinal luminal TNF-alpha levels were also measured. Production of hepatic IL-18 and TNF-alpha and numbers of KC were increased in rats with NEC and correlated with the progression of intestinal damage during NEC development. Furthermore, increased levels of TNF-alpha in the intestinal lumen of rats with NEC was significantly decreased when KC were inhibited with gadolinium ch...

Research paper thumbnail of Milk-Borne EGF and Necrotizing Enterocolitis in Neonatal Rat Model

Advances in Experimental Medicine and Biology, 2002

Neonatal necrotizing enterocolitis (NEC) is the most common gastrointestinal (GI) disease of prem... more Neonatal necrotizing enterocolitis (NEC) is the most common gastrointestinal (GI) disease of premature infants with excessive morbidity and mortality that afflicts 3,000 to 4,000 babies in the United States each year.1 Many factors contribute to the development of NEC, mainly prematurity, enteral feeding, infectious agents and/or intestinal hypoxia-ischemia. Enteral feeding is nearly always a prerequisite for the development of NEC, but the exact mechanism of NEC pathogenesis is poorly understood. The protective role of maternal milk in NEC pathogenesis has been reported.2 Various components of milk have been tested to protect the gut against NEC.3 Epidermal growth factor (EGF) is a promising candidate for the treatment of NEC. Mammalian milk of many species contains high concentrations of EGF. Moreover, maternal milk is the major source of EGF for neonates during the suckling period.4 In contrast, EGF is absent in all commercial infant formulas. Strong effects of exogenous EGF on healing of damaged gastrointestinal mucosa or on intestinal adaptation after injury are reported in a number of studies.5 The aim of this study was to examine the effects of milk-borne EGF on the development of NEC in a neonatal rat model.

Research paper thumbnail of Rat Milk Decreases Necrotizing Enterocolitis in a Rat Model

Advances in Experimental Medicine and Biology, 2004

... MILK DECREASES NECROTIZING ENTEROCOLITIS IN A RAT MODEL Bohuslav Dvorak1, 2, Melissa D. Halpe... more ... MILK DECREASES NECROTIZING ENTEROCOLITIS IN A RAT MODEL Bohuslav Dvorak1, 2, Melissa D. Halpem1, Hana Holubec3, Katerina Dvorakova3, Jessica ... and other essential intestinal functions, plays a protective role against this disease (Schanler 2001; Lucas & Cole ...

Research paper thumbnail of Mechanisms of Methicillin-Resistant Staphylococcus aureus Pneumonia–Induced Intestinal Epithelial Apoptosis

Shock, 2012

Methicillin-resistant Staphylococcus aureus (MRSA) pneumonia-induced sepsis is a common cause of ... more Methicillin-resistant Staphylococcus aureus (MRSA) pneumonia-induced sepsis is a common cause of morbidity in the intensive care unit. Although pneumonia is initiated in the lungs, extrapulmonary manifestations occur commonly. In light of the key role the intestine plays in the pathophysiology of sepsis, we sought to determine whether MRSA pneumonia induces intestinal injury. FVB/N mice were subjected to MRSA or sham pneumonia and sacrificed 24 hours later. Septic animals had a marked increase in intestinal epithelial apoptosis by both H&E and active caspase-3 staining. MRSA-induced intestinal apoptosis was associated with an increase in the expression of the pro-apoptotic proteins Bid and Bax and the anti-apoptotic protein Bcl-x L in the mitochondrial pathway. In the receptor-mediated pathway, MRSA pneumonia induced an increase in Fas-ligand but decreased protein levels of Fas, FADD, pFADD, TNF-R1 and TRADD. To assess the functional significance of these changes, MRSA pneumonia was induced in mice with genetic manipulations in proteins in either the mitochondrial or receptor-mediated pathways. Both Bid −/− mice and animals with intestine specific overexpression of Bcl-2 had decreased intestinal apoptosis compared to wild type animals. In contrast, Fas-ligand −/− mice had no alterations in apoptosis. To determine if these findings were organism-specific, similar experiments were performed in mice subjected to Pseudomonas aeruginosa pneumonia. P. aeruginosa induced gut apoptosis, but unlike MRSA, this was associated with increased Bcl-2 and TNF-R1 and decreased Fas. MRSA pneumonia thus induces organism-specific changes in intestinal apoptosis via changes in both the mitochondrial and receptor-mediated pathways although the former may be more functionally significant.

Research paper thumbnail of Epidermal Growth Factor Improves SurvivaL and Prevents Intestinal Injury in a Murine Model of Pseudomonas aeruginosa Pneumonia

Shock, 2011

Mortality from pneumonia is mediated, in part, through extrapulmonary causes. Epidermal growth fa... more Mortality from pneumonia is mediated, in part, through extrapulmonary causes. Epidermal growth factor (EGF) has broad cytoprotective effects, including potent restorative properties in the injured intestine. The purpose of this study was to determine the efficacy of EGF treatment following Pseudomonas aeruginosa pneumonia. FVB/N mice underwent intratracheal injection of either Pseudomonas aeruginosa or saline and were then randomized to receive either systemic EGF or vehicle beginning immediately or 24 hours after the onset of pneumonia. Systemic EGF decreased seven-day mortality from 65% to 10% when initiated immediately after the onset of pneumonia and to 27% when initiated 24 hours after the onset of pneumonia. Even though injury in pneumonia is initiated in the lungs, the survival advantage conferred by EGF was not associated with improvements in pulmonary pathology. In contrast, EGF prevented intestinal injury by reversing pneumonia-induced increases in intestinal epithelial apoptosis and decreases in intestinal proliferation and villus length. Systemic cytokines, kidney and liver function were unaffected by EGF therapy although EGF decreased pneumonia-induced splenocyte apoptosis. To determine whether the intestine was sufficient to account for extrapulmonary effects induced by EGF, a separate set of experiments were done using transgenic mice with enterocyte-specific overexpression of EGF (IFABP-EGF mice) which were compared to WT mice subjected to pneumonia. IFABP-EGF mice had improved survival compared to WT mice following pneumonia (50% vs. 28% respectively, p<0.05) and were protected from pneumonia-induced intestinal injury. Thus, EGF may be a potential adjunctive therapy for pneumonia, mediated in part by its effects on the intestine.

Research paper thumbnail of Cecal Ligation and Puncture Followed by Methicillin-Resistant Staphylococcus aureus Pneumonia Increases Mortality in Mice and Blunts Production of Local and Systemic Cytokines

Shock, 2012

Mortality in the intensive care unit frequently results from the synergistic effect of two tempor... more Mortality in the intensive care unit frequently results from the synergistic effect of two temporally distinct infections. This study examined the pathophysiology of a new model of intra-abdominal sepsis followed by methicillin-resistant Staphylococcus aureus (MRSA) pneumonia. Mice underwent cecal ligation and puncture (CLP) or sham laparotomy followed 3 days later by an intratracheal injection of MRSA or saline. Both CLP/saline and sham/MRSA mice had 100% survival, whereas animals with CLP followed by MRSA pneumonia had 67% 7-day survival. Animals subjected to CLP/MRSA had increased bronchoalveolar lavage concentrations of MRSA compared with sham/MRSA animals. Animals subjected to sham/MRSA pneumonia had increased bronchoalveolar lavage levels of interleukin 6 (IL-6), tumor necrosis factor α, and granulocyte colony-stimulating factor compared with those given intratracheal saline, whereas CLP/MRSA mice had a blunted local inflammatory response with markedly decreased cytokine levels. Similarly, animals subjected to CLP/saline had increased peritoneal lavage levels of IL-6 and IL-1β compared with those subjected to sham laparotomy, whereas this response was blunted in CLP/MRSA mice. Systemic cytokines were upregulated in both CLP/saline and sham/MRSA mice, and this was blunted by the combination of CLP/MRSA. In contrast, no synergistic effect on pneumonia severity, white blood cell count, or lymphocyte apoptosis was identified in CLP/MRSA mice compared with animals with either insult in isolation. These results indicate that a clinically relevant model of CLP followed by MRSA pneumonia causes higher mortality than could have been predicted from studying either infection in isolation, and this was associated with a blunted local (pulmonary and peritoneal) and systemic inflammatory response and decreased ability to clear infection.

Research paper thumbnail of Intestine-Specific Mttp Deletion Decreases Mortality and Prevents Sepsis-Induced Intestinal Injury in a Murine Model of Pseudomonas aeruginosa Pneumonia

PLoS ONE, 2012

Background: The small intestine plays a crucial role in the pathophysiology of sepsis and has bee... more Background: The small intestine plays a crucial role in the pathophysiology of sepsis and has been referred to as the ''motor'' of the systemic inflammatory response. One proposed mechanism is that toxic gut-derived lipid factors, transported in mesenteric lymph, induce systemic injury and distant organ failure. However, the pathways involved are yet to be defined and the role of intestinal chylomicron assembly and secretion in transporting these lipid factors is unknown. Here we studied the outcome of sepsis in mice with conditional, intestine-specific deletion of microsomal triglyceride transfer protein (Mttp-IKO), which exhibit a block in chylomicron assembly together with lipid malabsorption. Methodology/Principal Findings: Mttp-IKO mice and controls underwent intratracheal injection with either Pseudomonas aeruginosa or sterile saline. Mttp-IKO mice exhibited decreased seven-day mortality, with 0/20 (0%) dying compared to 5/17 (29%) control mice (p,0.05). This survival advantage in Mttp-IKO mice, however, was not associated with improvements in pulmonary bacterial clearance or neutrophil infiltration. Rather, Mttp-IKO mice exhibited protection against sepsisassociated decreases in villus length and intestinal proliferation and were also protected against increased intestinal apoptosis, both central features in control septic mice. Serum IL-6 levels, a major predictor of mortality in human and mouse models of sepsis, were elevated 8-fold in septic control mice but remained unaltered in septic Mttp-IKO mice. Serum high density lipoprotein (HDL) levels were reduced in septic control mice but were increased in septic Mttp-IKO mice. The decreased levels of HDL were associated with decreased hepatic expression of apolipoprotein A1 in septic control mice. Conclusions/Significance: These studies suggest that strategies directed at blocking intestinal chylomicron secretion may attenuate the progression and improve the outcome of sepsis through effects mediated by metabolic and physiological adaptations in both intestinal and hepatic lipid flux.

Research paper thumbnail of Maternal Milk Reduces Severity of Necrotizing Enterocolitis and Increases Intestinal IL-10 in a Neonatal Rat Model

Pediatric Research, 2003

Necrotizing enterocolitis (NEC) is a devastating intestinal disease of premature infants. Materna... more Necrotizing enterocolitis (NEC) is a devastating intestinal disease of premature infants. Maternal milk has been suggested to be partially protective against NEC; however, the mechanisms of this protection are not defined. The aim of this study was to examine the effect(s) of artificial feeding of rat milk (RM)versus cow milk-based rat milk substitute (RMS) on the development of NEC in a neonatal rat model and elucidate the role of inflammatory cytokines in NEC pathogenesis. Newborn rats were artificially fed with either collected RM or RMS. Experimental NEC was induced by exposure to asphyxia and cold stress and evaluated by histologic scoring of damage in ileum. Intestinal cytokine mRNA expression was determined by real-time PCR. Cytokine histologic localization was performed by confocal microscopy. Similar to human NEC, artificial feeding of RM reduces the incidence and severity of NEC injury in neonatal rats. Freezing and thawing of collected RM did not eliminate the protective effect of maternal milk. Ileal IL-10 expression was significantly increased in the RM group compared with RMS.

Research paper thumbnail of Milk-Borne Epidermal Growth Factor Modulates Intestinal Transforming Growth Factor-α Levels in Neonatal Rats

Research paper thumbnail of Up-Regulation of IL-18 and IL-12 in the Ileum of Neonatal Rats with Necrotizing Enterocolitis

Pediatric Research, 2002

Necrotizing enterocolitis (NEC) is a common and devastating gastrointestinal disease of premature... more Necrotizing enterocolitis (NEC) is a common and devastating gastrointestinal disease of premature infants. Because the proinflammatory cytokines IL-18, IL-12, and interferon (IFN)-␥ have been implicated in other diseases of the small intestine, we hypothesized that these cytokines would play an important role in NEC pathogenesis. NEC was induced in newborn rats via enteral feeding with rat milk substitute and asphyxia and cold stress (RMS). Dam-fed, asphyxia-and cold-stressed littermates were used as controls (DF). After 96 h, the distal ileum was removed from all animals and processed to determine expression and localization of IL-18, IL-12, and IFN-␥ using real-time reverse transcriptase PCR and immunohistology. IL-18 and IL-12 mRNA from the RMS group were increased (p Յ 0.05) compared with DF controls, and there was a correlation between increasing IL-18 and IL-12 mRNA levels and progression of tissue damage (r ϭ 0.629 and 0.588, respectively; p Յ 0.05). Immunohistology revealed IL-18 in the cytoplasm of villi and crypt enterocytes and IL-12-positive monocytes/macrophages were increased with disease progression (r ϭ 0.503, p Յ 0.05). No differences in the number of IFN-␥-positive cells were observed between groups. These data demonstrate up-regulation of IL-18 and IL-12 in experimental NEC and a correlation between production of these proinflammatory cytokines and progression of tissue damage. (Pediatr Res 51: 733-739, 2002) Abbreviations DF, dam fed IEC, intestinal epithelial cells IFN-␥, interferon-␥ LP, lamina propria LPMC, lamina propria monocytes/macrophages MC, monocytes/macrophages NEC, necrotizing enterocolitis NLPMC, non-lamina propria monocytes/macrophages RT-PCR, reverse transcriptase PCR RMS, rat milk substitute

Research paper thumbnail of Cecal ligation and puncture followed by MRSA pneumonia increases mortality in mice and blunts production of local and systemic cytokines

Journal of the American College of Surgeons, 2011

Mortality in the ICU frequently results from the synergistic effect of two temporally-distinct in... more Mortality in the ICU frequently results from the synergistic effect of two temporally-distinct infections. This study examined the pathophysiology of a new model of intraabdominal sepsis followed by methicillin-resistant Staphylococcus aureus (MRSA) pneumonia. Mice underwent cecal ligation and puncture (CLP) or sham laparotomy followed three days later by an intratracheal injection of MRSA or saline. Both CLP/saline and sham/MRSA mice had 100% survival while animals with CLP followed by MRSA pneumonia had 67% seven-day survival. Animals subjected to CLP/MRSA had increased bronchoalveolar lavage (BAL) concentrations of MRSA compared to sham/MRSA animals. Animals subjected to sham/MRSA pneumonia had increased BAL levels of IL-6, TNF-α, and G-CSF compared to those given intratracheal saline while CLP/MRSA mice had a blunted local inflammatory response with markedly decreased cytokine levels. Similarly, animals subjected to CLP/saline had increased peritoneal lavage levels of IL-6 and IL-1β compared to those subjected to sham laparotomy while this response was blunted in CLP/MRSA mice. Systemic cytokines were upregulated in both CLP/saline and sham/ MRSA mice, and this was blunted by the combination of CLP/MRSA. In contrast, no synergistic effect on pneumonia severity, white blood cell count or lymphocyte apoptosis was identified in CLP/MRSA mice compared to animals with either insult in isolation. These results indicate that a clinically relevant model of CLP followed by MRSA pneumonia causes higher mortality than

Research paper thumbnail of Ileal Cytokine Dysregulation in Experimental Necrotizing Enterocolitis Is Reduced by Epidermal Growth Factor

Journal of Pediatric Gastroenterology and Nutrition, 2003

Background: Necrotizing enterocolitis (NEC) is the most common gastrointestinal disease of premat... more Background: Necrotizing enterocolitis (NEC) is the most common gastrointestinal disease of premature infants. We have shown in previous studies that proinflammatory interleukin-18 and interleukin-12 are up-regulated in the ileum of rats with experimental NEC and that epidermal growth factor (EGF) reduces the development of disease. Here we investigated whether the protective effects of EGF are a result of changes in ileal interleukin-18, interleukin-12 and/or antiinflammatory interleukin-10. Methods: Newborn rats were artificially fed with either growth-factor-free rat milk substitute (RMS) or RMS supplemented with 500 ng/mL EGF (RMS + EGF) and NEC was induced via exposure to asphyxia and cold stress. Cytokine expression and localization were assessed using reverse-transcription real-time polymerase chain reaction and immunohistology/confocal microscopy. Results: Enteral administration of EGF (RMS + EGF) decreased overproduction of interleukin-18 and increased interleukin-10 production in the ileum. Furthermore, increased interleukin-10 production was associated with up-regulation of the transcription factor Sp1 in RMS + EGF rats. Conclusions: These data suggest that EGF may reduce NEC via increased interleukin-10 and decreased interleukin-18 and that EGF-mediated up-regulation of Sp1 may account for the increased interleukin-10. JPGN 36:126-133, 2003.

Research paper thumbnail of Artificial Formula Induces Precocious Maturation of the Small Intestine of Artificially Reared Suckling Rats

Journal of Pediatric Gastroenterology and Nutrition, 2000

Background: The artificially reared rat model was used successfully to study the effect of nutrit... more Background: The artificially reared rat model was used successfully to study the effect of nutrition during the early postnatal period on growth and development of the neonate. Overgrowth and morphologic changes of the gastrointestinal tract are known consequences of artificial rearing. The major goal of our study was to elucidate whether artificial rearing-enhanced gut development is caused by artificial diet or by gastrostomy and the artificial rearing technique itself. Methods: Suckling rats at day 8 of age underwent intragastric cannulation and were machine fed either a cow's milk-based artificial rat's milk substitute or pooled rat's milk for 4 days. Dam-fed littermates served as a control. Results: Body growth did not differ in the three experimental groups. In rats receiving rat's milk substitute, small intestinal wet weight was approximately 60% greater than in rats fed rat's milk or control rats. Additionally, the entire small intestine was approximately 20% longer in the rat's milk substitute group. Morphologically, rat's milk substitute-fed pups demonstrated significantly greater intestinal villus length and crypt depth compared with rat's milk-fed or control rats. Jejunum and midjejunum of the rat's milk and control groups did not differ in these parameters. Intestinal sucrase activity of rat's milk substitute-fed rats was significantly elevated compared with rat's milk-fed rats or control animals. Conclusions: These results indicate that cow's milk-based formula, not gastrostomy or artificial feeding technique, is a principal cause of the small intestine overgrowth and precocious maturation of some intestinal functions observed in artificially reared sucklings.

Research paper thumbnail of Prevention of Lymphocyte Apoptosis in Septic Mice with Cancer Increases Mortality

The Journal of Immunology, 2011

Lymphocyte apoptosis is thought to have a major role in the pathophysiology of sepsis. However, t... more Lymphocyte apoptosis is thought to have a major role in the pathophysiology of sepsis. However, there is a disconnect between animal models of sepsis and patients with the disease, because the former use subjects that were healthy prior to the onset of infection while most patients have underlying comorbidities. The purpose of this study was to determine whether lymphocyte apoptosis prevention is effective in preventing mortality in septic mice with preexisting cancer. Mice with lymphocyte Bcl-2 overexpression (Bcl-2-Ig) and wild type (WT) mice were injected with a transplantable pancreatic adenocarcinoma cell line. Three weeks later, after development of palpable tumors, all animals received an intratracheal injection of Pseudomonas aeruginosa. Despite having decreased sepsis-induced T and B lymphocyte apoptosis, Bcl-2-Ig mice had markedly increased mortality compared with WT mice following P. aeruginosa pneumonia (85 versus 44% 7-d mortality; p = 0.004). The worsened survival in B...

Research paper thumbnail of TNF is a key mediator in sepsis‐induced intestinal barrier dysfunction but is independent of enterocyte NFκB

Research paper thumbnail of Epidermal growth factor treatment prevents intestinal injury in weanling mice with septic peritonitis

Research paper thumbnail of Plan de empresa Jeshia Construction

Según la demanda de proyectos solicitados contrataremos a más profesionales del sector de la cons... more Según la demanda de proyectos solicitados contrataremos a más profesionales del sector de la construcción.

Research paper thumbnail of Plan De Relaciones Publicas Para Agencia De Investigacion De Mercados

Research paper thumbnail of Intestine-Specific Deletion of Microsomal Triglyceride Transfer Protein Increases Mortality in Aged Mice

PLoS ONE, 2014

Background: Mice with conditional, intestine-specific deletion of microsomal triglyceride transfe... more Background: Mice with conditional, intestine-specific deletion of microsomal triglyceride transfer protein (Mttp-IKO) exhibit a complete block in chylomicron assembly together with lipid malabsorption. Young (8-10 week) Mttp-IKO mice have improved survival when subjected to a murine model of Pseudomonas aeruginosa-induced sepsis. However, 80% of deaths in sepsis occur in patients over age 65. The purpose of this study was to determine whether age impacts outcome in Mttp-IKO mice subjected to sepsis. Methods: Aged (20-24 months) Mttp-IKO mice and WT mice underwent intratracheal injection with P. aeruginosa. Mice were either sacrificed 24 hours post-operatively for mechanistic studies or followed seven days for survival. Results: In contrast to young septic Mttp-IKO mice, aged septic Mttp-IKO mice had a significantly higher mortality than aged septic WT mice (80% vs. 39%, p = 0.005). Aged septic Mttp-IKO mice exhibited increased gut epithelial apoptosis, increased jejunal Bax/Bcl-2 and Bax/Bcl-X L ratios yet simultaneously demonstrated increased crypt proliferation and villus length. Aged septic Mttp-IKO mice also manifested increased pulmonary myeloperoxidase levels, suggesting increased neutrophil infiltration, as well as decreased systemic TNFa compared to aged septic WT mice. Conclusions: Blocking intestinal chylomicron secretion alters mortality following sepsis in an age-dependent manner. Increases in gut apoptosis and pulmonary neutrophil infiltration, and decreased systemic TNFa represent potential mechanisms for why intestine-specific Mttp deletion is beneficial in young septic mice but harmful in aged mice as each of these parameters are altered differently in young and aged septic WT and Mttp-IKO mice.

Research paper thumbnail of Epidermal growth factor reduces the development of necrotizing enterocolitis in a neonatal rat model

American journal of physiology. Gastrointestinal and liver physiology, 2002

Necrotizing enterocolitis (NEC) is the most common gastrointestinal disease of prematurely born i... more Necrotizing enterocolitis (NEC) is the most common gastrointestinal disease of prematurely born infants. Maternal milk plays an important protective role against NEC development and is the major source of epidermal growth factor (EGF) for neonates. The aim of this study was to examine the effect of orally administered EGF on the incidence of NEC in a neonatal rat model. Newborn rats were artificially fed either with growth factor-free rat milk substitute (RMS) or RMS supplemented with 500 ng/ml of EGF (RMS+EGF). Experimental NEC was induced by exposure to asphyxia and cold stress. Development of NEC was evaluated by gross and histological scoring of damage in the ileum. Ileal EGF receptor (EGF-R), EGF, and transforming growth factor-alpha mRNA expression was assessed by RT competitive-PCR, and the EGF-R was localized by immunohistochemistry. EGF supplementation of formula reduced the incidence and severity of NEC in rats (13/16 RMS vs. 4/13 RMS+EGF). Ileal EGF-R mRNA expression was ...

Research paper thumbnail of Hepatic inflammatory mediators contribute to intestinal damage in necrotizing enterocolitis

American journal of physiology. Gastrointestinal and liver physiology, 2003

Necrotizing enterocolitis (NEC) is a common and devastating gastrointestinal disease of premature... more Necrotizing enterocolitis (NEC) is a common and devastating gastrointestinal disease of premature infants. Along with pathological effects in the ileum, severe NEC is often accompanied by multisystem organ failure, including liver failure. The aim of this study was to determine the changes in hepatic cytokines and inflammatory mediators in experimental NEC. The well-established neonatal rat model of NEC was used in this study, and changes in liver morphology, numbers of Kupffer cells (KC), gene expression, and histological localization of IL-18, TNF-alpha, and inducible nitric oxide synthase were evaluated. Intestinal luminal TNF-alpha levels were also measured. Production of hepatic IL-18 and TNF-alpha and numbers of KC were increased in rats with NEC and correlated with the progression of intestinal damage during NEC development. Furthermore, increased levels of TNF-alpha in the intestinal lumen of rats with NEC was significantly decreased when KC were inhibited with gadolinium ch...

Research paper thumbnail of Milk-Borne EGF and Necrotizing Enterocolitis in Neonatal Rat Model

Advances in Experimental Medicine and Biology, 2002

Neonatal necrotizing enterocolitis (NEC) is the most common gastrointestinal (GI) disease of prem... more Neonatal necrotizing enterocolitis (NEC) is the most common gastrointestinal (GI) disease of premature infants with excessive morbidity and mortality that afflicts 3,000 to 4,000 babies in the United States each year.1 Many factors contribute to the development of NEC, mainly prematurity, enteral feeding, infectious agents and/or intestinal hypoxia-ischemia. Enteral feeding is nearly always a prerequisite for the development of NEC, but the exact mechanism of NEC pathogenesis is poorly understood. The protective role of maternal milk in NEC pathogenesis has been reported.2 Various components of milk have been tested to protect the gut against NEC.3 Epidermal growth factor (EGF) is a promising candidate for the treatment of NEC. Mammalian milk of many species contains high concentrations of EGF. Moreover, maternal milk is the major source of EGF for neonates during the suckling period.4 In contrast, EGF is absent in all commercial infant formulas. Strong effects of exogenous EGF on healing of damaged gastrointestinal mucosa or on intestinal adaptation after injury are reported in a number of studies.5 The aim of this study was to examine the effects of milk-borne EGF on the development of NEC in a neonatal rat model.

Research paper thumbnail of Rat Milk Decreases Necrotizing Enterocolitis in a Rat Model

Advances in Experimental Medicine and Biology, 2004

... MILK DECREASES NECROTIZING ENTEROCOLITIS IN A RAT MODEL Bohuslav Dvorak1, 2, Melissa D. Halpe... more ... MILK DECREASES NECROTIZING ENTEROCOLITIS IN A RAT MODEL Bohuslav Dvorak1, 2, Melissa D. Halpem1, Hana Holubec3, Katerina Dvorakova3, Jessica ... and other essential intestinal functions, plays a protective role against this disease (Schanler 2001; Lucas &amp;amp; Cole ...

Research paper thumbnail of Mechanisms of Methicillin-Resistant Staphylococcus aureus Pneumonia–Induced Intestinal Epithelial Apoptosis

Shock, 2012

Methicillin-resistant Staphylococcus aureus (MRSA) pneumonia-induced sepsis is a common cause of ... more Methicillin-resistant Staphylococcus aureus (MRSA) pneumonia-induced sepsis is a common cause of morbidity in the intensive care unit. Although pneumonia is initiated in the lungs, extrapulmonary manifestations occur commonly. In light of the key role the intestine plays in the pathophysiology of sepsis, we sought to determine whether MRSA pneumonia induces intestinal injury. FVB/N mice were subjected to MRSA or sham pneumonia and sacrificed 24 hours later. Septic animals had a marked increase in intestinal epithelial apoptosis by both H&E and active caspase-3 staining. MRSA-induced intestinal apoptosis was associated with an increase in the expression of the pro-apoptotic proteins Bid and Bax and the anti-apoptotic protein Bcl-x L in the mitochondrial pathway. In the receptor-mediated pathway, MRSA pneumonia induced an increase in Fas-ligand but decreased protein levels of Fas, FADD, pFADD, TNF-R1 and TRADD. To assess the functional significance of these changes, MRSA pneumonia was induced in mice with genetic manipulations in proteins in either the mitochondrial or receptor-mediated pathways. Both Bid −/− mice and animals with intestine specific overexpression of Bcl-2 had decreased intestinal apoptosis compared to wild type animals. In contrast, Fas-ligand −/− mice had no alterations in apoptosis. To determine if these findings were organism-specific, similar experiments were performed in mice subjected to Pseudomonas aeruginosa pneumonia. P. aeruginosa induced gut apoptosis, but unlike MRSA, this was associated with increased Bcl-2 and TNF-R1 and decreased Fas. MRSA pneumonia thus induces organism-specific changes in intestinal apoptosis via changes in both the mitochondrial and receptor-mediated pathways although the former may be more functionally significant.

Research paper thumbnail of Epidermal Growth Factor Improves SurvivaL and Prevents Intestinal Injury in a Murine Model of Pseudomonas aeruginosa Pneumonia

Shock, 2011

Mortality from pneumonia is mediated, in part, through extrapulmonary causes. Epidermal growth fa... more Mortality from pneumonia is mediated, in part, through extrapulmonary causes. Epidermal growth factor (EGF) has broad cytoprotective effects, including potent restorative properties in the injured intestine. The purpose of this study was to determine the efficacy of EGF treatment following Pseudomonas aeruginosa pneumonia. FVB/N mice underwent intratracheal injection of either Pseudomonas aeruginosa or saline and were then randomized to receive either systemic EGF or vehicle beginning immediately or 24 hours after the onset of pneumonia. Systemic EGF decreased seven-day mortality from 65% to 10% when initiated immediately after the onset of pneumonia and to 27% when initiated 24 hours after the onset of pneumonia. Even though injury in pneumonia is initiated in the lungs, the survival advantage conferred by EGF was not associated with improvements in pulmonary pathology. In contrast, EGF prevented intestinal injury by reversing pneumonia-induced increases in intestinal epithelial apoptosis and decreases in intestinal proliferation and villus length. Systemic cytokines, kidney and liver function were unaffected by EGF therapy although EGF decreased pneumonia-induced splenocyte apoptosis. To determine whether the intestine was sufficient to account for extrapulmonary effects induced by EGF, a separate set of experiments were done using transgenic mice with enterocyte-specific overexpression of EGF (IFABP-EGF mice) which were compared to WT mice subjected to pneumonia. IFABP-EGF mice had improved survival compared to WT mice following pneumonia (50% vs. 28% respectively, p<0.05) and were protected from pneumonia-induced intestinal injury. Thus, EGF may be a potential adjunctive therapy for pneumonia, mediated in part by its effects on the intestine.

Research paper thumbnail of Cecal Ligation and Puncture Followed by Methicillin-Resistant Staphylococcus aureus Pneumonia Increases Mortality in Mice and Blunts Production of Local and Systemic Cytokines

Shock, 2012

Mortality in the intensive care unit frequently results from the synergistic effect of two tempor... more Mortality in the intensive care unit frequently results from the synergistic effect of two temporally distinct infections. This study examined the pathophysiology of a new model of intra-abdominal sepsis followed by methicillin-resistant Staphylococcus aureus (MRSA) pneumonia. Mice underwent cecal ligation and puncture (CLP) or sham laparotomy followed 3 days later by an intratracheal injection of MRSA or saline. Both CLP/saline and sham/MRSA mice had 100% survival, whereas animals with CLP followed by MRSA pneumonia had 67% 7-day survival. Animals subjected to CLP/MRSA had increased bronchoalveolar lavage concentrations of MRSA compared with sham/MRSA animals. Animals subjected to sham/MRSA pneumonia had increased bronchoalveolar lavage levels of interleukin 6 (IL-6), tumor necrosis factor α, and granulocyte colony-stimulating factor compared with those given intratracheal saline, whereas CLP/MRSA mice had a blunted local inflammatory response with markedly decreased cytokine levels. Similarly, animals subjected to CLP/saline had increased peritoneal lavage levels of IL-6 and IL-1β compared with those subjected to sham laparotomy, whereas this response was blunted in CLP/MRSA mice. Systemic cytokines were upregulated in both CLP/saline and sham/MRSA mice, and this was blunted by the combination of CLP/MRSA. In contrast, no synergistic effect on pneumonia severity, white blood cell count, or lymphocyte apoptosis was identified in CLP/MRSA mice compared with animals with either insult in isolation. These results indicate that a clinically relevant model of CLP followed by MRSA pneumonia causes higher mortality than could have been predicted from studying either infection in isolation, and this was associated with a blunted local (pulmonary and peritoneal) and systemic inflammatory response and decreased ability to clear infection.

Research paper thumbnail of Intestine-Specific Mttp Deletion Decreases Mortality and Prevents Sepsis-Induced Intestinal Injury in a Murine Model of Pseudomonas aeruginosa Pneumonia

PLoS ONE, 2012

Background: The small intestine plays a crucial role in the pathophysiology of sepsis and has bee... more Background: The small intestine plays a crucial role in the pathophysiology of sepsis and has been referred to as the ''motor'' of the systemic inflammatory response. One proposed mechanism is that toxic gut-derived lipid factors, transported in mesenteric lymph, induce systemic injury and distant organ failure. However, the pathways involved are yet to be defined and the role of intestinal chylomicron assembly and secretion in transporting these lipid factors is unknown. Here we studied the outcome of sepsis in mice with conditional, intestine-specific deletion of microsomal triglyceride transfer protein (Mttp-IKO), which exhibit a block in chylomicron assembly together with lipid malabsorption. Methodology/Principal Findings: Mttp-IKO mice and controls underwent intratracheal injection with either Pseudomonas aeruginosa or sterile saline. Mttp-IKO mice exhibited decreased seven-day mortality, with 0/20 (0%) dying compared to 5/17 (29%) control mice (p,0.05). This survival advantage in Mttp-IKO mice, however, was not associated with improvements in pulmonary bacterial clearance or neutrophil infiltration. Rather, Mttp-IKO mice exhibited protection against sepsisassociated decreases in villus length and intestinal proliferation and were also protected against increased intestinal apoptosis, both central features in control septic mice. Serum IL-6 levels, a major predictor of mortality in human and mouse models of sepsis, were elevated 8-fold in septic control mice but remained unaltered in septic Mttp-IKO mice. Serum high density lipoprotein (HDL) levels were reduced in septic control mice but were increased in septic Mttp-IKO mice. The decreased levels of HDL were associated with decreased hepatic expression of apolipoprotein A1 in septic control mice. Conclusions/Significance: These studies suggest that strategies directed at blocking intestinal chylomicron secretion may attenuate the progression and improve the outcome of sepsis through effects mediated by metabolic and physiological adaptations in both intestinal and hepatic lipid flux.

Research paper thumbnail of Maternal Milk Reduces Severity of Necrotizing Enterocolitis and Increases Intestinal IL-10 in a Neonatal Rat Model

Pediatric Research, 2003

Necrotizing enterocolitis (NEC) is a devastating intestinal disease of premature infants. Materna... more Necrotizing enterocolitis (NEC) is a devastating intestinal disease of premature infants. Maternal milk has been suggested to be partially protective against NEC; however, the mechanisms of this protection are not defined. The aim of this study was to examine the effect(s) of artificial feeding of rat milk (RM)versus cow milk-based rat milk substitute (RMS) on the development of NEC in a neonatal rat model and elucidate the role of inflammatory cytokines in NEC pathogenesis. Newborn rats were artificially fed with either collected RM or RMS. Experimental NEC was induced by exposure to asphyxia and cold stress and evaluated by histologic scoring of damage in ileum. Intestinal cytokine mRNA expression was determined by real-time PCR. Cytokine histologic localization was performed by confocal microscopy. Similar to human NEC, artificial feeding of RM reduces the incidence and severity of NEC injury in neonatal rats. Freezing and thawing of collected RM did not eliminate the protective effect of maternal milk. Ileal IL-10 expression was significantly increased in the RM group compared with RMS.

Research paper thumbnail of Milk-Borne Epidermal Growth Factor Modulates Intestinal Transforming Growth Factor-α Levels in Neonatal Rats

Research paper thumbnail of Up-Regulation of IL-18 and IL-12 in the Ileum of Neonatal Rats with Necrotizing Enterocolitis

Pediatric Research, 2002

Necrotizing enterocolitis (NEC) is a common and devastating gastrointestinal disease of premature... more Necrotizing enterocolitis (NEC) is a common and devastating gastrointestinal disease of premature infants. Because the proinflammatory cytokines IL-18, IL-12, and interferon (IFN)-␥ have been implicated in other diseases of the small intestine, we hypothesized that these cytokines would play an important role in NEC pathogenesis. NEC was induced in newborn rats via enteral feeding with rat milk substitute and asphyxia and cold stress (RMS). Dam-fed, asphyxia-and cold-stressed littermates were used as controls (DF). After 96 h, the distal ileum was removed from all animals and processed to determine expression and localization of IL-18, IL-12, and IFN-␥ using real-time reverse transcriptase PCR and immunohistology. IL-18 and IL-12 mRNA from the RMS group were increased (p Յ 0.05) compared with DF controls, and there was a correlation between increasing IL-18 and IL-12 mRNA levels and progression of tissue damage (r ϭ 0.629 and 0.588, respectively; p Յ 0.05). Immunohistology revealed IL-18 in the cytoplasm of villi and crypt enterocytes and IL-12-positive monocytes/macrophages were increased with disease progression (r ϭ 0.503, p Յ 0.05). No differences in the number of IFN-␥-positive cells were observed between groups. These data demonstrate up-regulation of IL-18 and IL-12 in experimental NEC and a correlation between production of these proinflammatory cytokines and progression of tissue damage. (Pediatr Res 51: 733-739, 2002) Abbreviations DF, dam fed IEC, intestinal epithelial cells IFN-␥, interferon-␥ LP, lamina propria LPMC, lamina propria monocytes/macrophages MC, monocytes/macrophages NEC, necrotizing enterocolitis NLPMC, non-lamina propria monocytes/macrophages RT-PCR, reverse transcriptase PCR RMS, rat milk substitute

Research paper thumbnail of Cecal ligation and puncture followed by MRSA pneumonia increases mortality in mice and blunts production of local and systemic cytokines

Journal of the American College of Surgeons, 2011

Mortality in the ICU frequently results from the synergistic effect of two temporally-distinct in... more Mortality in the ICU frequently results from the synergistic effect of two temporally-distinct infections. This study examined the pathophysiology of a new model of intraabdominal sepsis followed by methicillin-resistant Staphylococcus aureus (MRSA) pneumonia. Mice underwent cecal ligation and puncture (CLP) or sham laparotomy followed three days later by an intratracheal injection of MRSA or saline. Both CLP/saline and sham/MRSA mice had 100% survival while animals with CLP followed by MRSA pneumonia had 67% seven-day survival. Animals subjected to CLP/MRSA had increased bronchoalveolar lavage (BAL) concentrations of MRSA compared to sham/MRSA animals. Animals subjected to sham/MRSA pneumonia had increased BAL levels of IL-6, TNF-α, and G-CSF compared to those given intratracheal saline while CLP/MRSA mice had a blunted local inflammatory response with markedly decreased cytokine levels. Similarly, animals subjected to CLP/saline had increased peritoneal lavage levels of IL-6 and IL-1β compared to those subjected to sham laparotomy while this response was blunted in CLP/MRSA mice. Systemic cytokines were upregulated in both CLP/saline and sham/ MRSA mice, and this was blunted by the combination of CLP/MRSA. In contrast, no synergistic effect on pneumonia severity, white blood cell count or lymphocyte apoptosis was identified in CLP/MRSA mice compared to animals with either insult in isolation. These results indicate that a clinically relevant model of CLP followed by MRSA pneumonia causes higher mortality than

Research paper thumbnail of Ileal Cytokine Dysregulation in Experimental Necrotizing Enterocolitis Is Reduced by Epidermal Growth Factor

Journal of Pediatric Gastroenterology and Nutrition, 2003

Background: Necrotizing enterocolitis (NEC) is the most common gastrointestinal disease of premat... more Background: Necrotizing enterocolitis (NEC) is the most common gastrointestinal disease of premature infants. We have shown in previous studies that proinflammatory interleukin-18 and interleukin-12 are up-regulated in the ileum of rats with experimental NEC and that epidermal growth factor (EGF) reduces the development of disease. Here we investigated whether the protective effects of EGF are a result of changes in ileal interleukin-18, interleukin-12 and/or antiinflammatory interleukin-10. Methods: Newborn rats were artificially fed with either growth-factor-free rat milk substitute (RMS) or RMS supplemented with 500 ng/mL EGF (RMS + EGF) and NEC was induced via exposure to asphyxia and cold stress. Cytokine expression and localization were assessed using reverse-transcription real-time polymerase chain reaction and immunohistology/confocal microscopy. Results: Enteral administration of EGF (RMS + EGF) decreased overproduction of interleukin-18 and increased interleukin-10 production in the ileum. Furthermore, increased interleukin-10 production was associated with up-regulation of the transcription factor Sp1 in RMS + EGF rats. Conclusions: These data suggest that EGF may reduce NEC via increased interleukin-10 and decreased interleukin-18 and that EGF-mediated up-regulation of Sp1 may account for the increased interleukin-10. JPGN 36:126-133, 2003.

Research paper thumbnail of Artificial Formula Induces Precocious Maturation of the Small Intestine of Artificially Reared Suckling Rats

Journal of Pediatric Gastroenterology and Nutrition, 2000

Background: The artificially reared rat model was used successfully to study the effect of nutrit... more Background: The artificially reared rat model was used successfully to study the effect of nutrition during the early postnatal period on growth and development of the neonate. Overgrowth and morphologic changes of the gastrointestinal tract are known consequences of artificial rearing. The major goal of our study was to elucidate whether artificial rearing-enhanced gut development is caused by artificial diet or by gastrostomy and the artificial rearing technique itself. Methods: Suckling rats at day 8 of age underwent intragastric cannulation and were machine fed either a cow's milk-based artificial rat's milk substitute or pooled rat's milk for 4 days. Dam-fed littermates served as a control. Results: Body growth did not differ in the three experimental groups. In rats receiving rat's milk substitute, small intestinal wet weight was approximately 60% greater than in rats fed rat's milk or control rats. Additionally, the entire small intestine was approximately 20% longer in the rat's milk substitute group. Morphologically, rat's milk substitute-fed pups demonstrated significantly greater intestinal villus length and crypt depth compared with rat's milk-fed or control rats. Jejunum and midjejunum of the rat's milk and control groups did not differ in these parameters. Intestinal sucrase activity of rat's milk substitute-fed rats was significantly elevated compared with rat's milk-fed rats or control animals. Conclusions: These results indicate that cow's milk-based formula, not gastrostomy or artificial feeding technique, is a principal cause of the small intestine overgrowth and precocious maturation of some intestinal functions observed in artificially reared sucklings.

Research paper thumbnail of Prevention of Lymphocyte Apoptosis in Septic Mice with Cancer Increases Mortality

The Journal of Immunology, 2011

Lymphocyte apoptosis is thought to have a major role in the pathophysiology of sepsis. However, t... more Lymphocyte apoptosis is thought to have a major role in the pathophysiology of sepsis. However, there is a disconnect between animal models of sepsis and patients with the disease, because the former use subjects that were healthy prior to the onset of infection while most patients have underlying comorbidities. The purpose of this study was to determine whether lymphocyte apoptosis prevention is effective in preventing mortality in septic mice with preexisting cancer. Mice with lymphocyte Bcl-2 overexpression (Bcl-2-Ig) and wild type (WT) mice were injected with a transplantable pancreatic adenocarcinoma cell line. Three weeks later, after development of palpable tumors, all animals received an intratracheal injection of Pseudomonas aeruginosa. Despite having decreased sepsis-induced T and B lymphocyte apoptosis, Bcl-2-Ig mice had markedly increased mortality compared with WT mice following P. aeruginosa pneumonia (85 versus 44% 7-d mortality; p = 0.004). The worsened survival in B...