Farokh R. Demehri

Intestinal epithelial cell apoptosis and loss of barrier function in the setting of altered microbiota with enteral nutrient deprivation

Total parenteral nutrition (TPN), a commonly used treatment for patients who cannot receive enteral nutrition, is associated with significant septic complications due in part to a loss of epithelial barrier function (EBF). While the underlying mechanisms of TPN-related epithelial changes are poorly understood, a mouse model of TPN-dependence has helped identify several contributing factors. Enteral deprivation leads to a shift in intestinal microbiota to predominantly Gram-negative Proteobacteria. This is associated with an increase in expression of proinflammatory cytokines within the mucosa, including interferon-γ and tumor necrosis factor-α. A concomitant loss of epithelial growth factors leads to a decrease in epithelial cell proliferation and increased apoptosis. The resulting loss of epithelial tight junction proteins contributes to EBF dysfunction. These mechanisms identify potential strategies of protecting against TPN-related complications, such as modification of luminal bacteria, blockade of proinflammatory cytokines, or growth factor replacement.

Hirschsprung-associated enterocolitis: pathogenesis, treatment and prevention

Hirschsprung-associated enterocolitis (HAEC) is a common and sometimes life-threatening complication of Hirschsprung disease (HD). Presenting either before or after definitive surgery for HD, HAEC may manifest clinically as abdominal distension and explosive diarrhea, along with emesis, fever, lethargy, and even shock. The pathogenesis of HAEC, the subject of ongoing research, likely involves a complex interplay between a dysfunctional enteric nervous system, abnormal mucin production, insufficient immunoglobulin secretion, and unbalanced intestinal microflora. Early recognition of HAEC and preventative practices, such as rectal washouts following a pull-through, can lead to improved outcomes. Treatment strategies for acute HAEC include timely resuscitation, colonic decompression, and antibiotics. Recurrent or persistent HAEC requires evaluation for mechanical obstruction or residual aganglionosis, and may require surgical treatment with posterior myotomy/myectomy or redo pull-through. This chapter describes the incidence, pathogenesis, treatment, and preventative strategies in management of HAEC.

Enteral nutrient deprivation in patients leads to a loss of intestinal epithelial barrier function

OBJECTIVE To investigate the effect of nutrient withdrawal on human intestinal epithelial barrier function (EBF). We hypothesized that unfed mucosa results in decreased EBF. This was tested in a series of surgical small intestinal resection specimens. DESIGN Small bowel specifically excluding inflamed tissue, was obtained from pediatric patients (aged 2 days to 19 years) undergoing intestinal resection. EBF was assessed in Ussing chambers for transepithelial resistance (TER) and passage of fluorescein isothiocyanate (FITC)-dextran (4 kD). Tight junction and adherence junction proteins were imaged with immunofluorescence staining. Expression of Toll-like receptors (TLR) and inflammatory cytokines were measured in loop ileostomy takedowns in a second group of patients. RESULTS Because TER increased with patient age (P < .01), results were stratified into infant versus teenage groups. Fed bowel had significantly greater TER versus unfed bowel (P < .05) in both age populations. Loss of EBF was also observed by an increase in FITC-dextran permeation in enteral nutrient-deprived segments (P < .05). Immunofluorescence staining showed marked declines in intensity of ZO-1, occludin, E-cadherin, and claudin-4 in unfed intestinal segments, as well as a loss of structural formation of tight junctions. Analysis of cytokine and TLR expression showed significant increases in tumor necrosis factor (TNF)-α and TLR4 in unfed segments of bowel compared with fed segments from the same individual. CONCLUSION EBF declined in unfed segments of human small bowel. This work represents the first direct examination of EBF from small bowel derived from nutrient-deprived humans and may explain the increased incidence of infectious complications seen in patients not receiving enteral feeds.

Enteral autonomy in pediatric short bowel syndrome: predictive factors one year after diagnosis

PURPOSE This study examined predictors of achieving enteral autonomy among pediatric short bowel syndrome (SBS) patients remaining on parenteral nutrition (PN) beyond one year. METHODS A retrospective single-institution study of 171 pediatric SBS patients (defined as ≥50% small bowel (SB) loss or ≥60 days of PN with onset before 6 weeks of age) was performed. Multivariate Cox proportional hazards analysis was conducted, with subgroup analysis of patients on PN for ≥1 year (n=59). Primary outcome was successful wean from PN. RESULTS Over a follow-up of 4.1±4.8 years, 64.3% of children weaned from PN. Mortality was 15.2%. Presence of ≥10% expected SB length (hazard ratio [HR] 6.48, p=0.002) or an ileocecal valve (ICV; HR, 2.86, p<0.001) predicted PN weaning. Of those on PN ≥1 year, the wean rate was 50.8%, and ICV no longer predicted weaning (p=0.153). Predictors among those on PN ≥1 year were: ≥10% expected SB length (HR, 8.27, p=0.010), intestinal atresia (HR, 4.26, p=0.011), and necrotizing enterocolitis (NEC, HR, 2.84, p=0.025). CONCLUSIONS SBS children on PN ≥1 year continue to wean from PN, and those with ≥10% of predicted SB length, NEC, or atresia are more likely to do so. These findings may help direct management and advice for these challenging patients.

Changes to the intestinal microbiome with parenteral nutrition: Review of a murine model and potential clinical implications

Parenteral nutrition (PN) dependence, while life sustaining, carries a significant risk of septic complications associated with epithelial barrier dysfunction and translocation of gut-derived microbiota. Increasing evidence suggests that PN-associated changes in the intestinal microbiota play a central role in the breakdown of the intestinal epithelial barrier. This review outlines the clinical and experimental evidence of epithelial barrier dysfunction with PN, the role of gut inflammatory dysregulation in driving this process, and the role of the intestinal microbiome in modulating inflammation in the gut and systemically. The article summarizes the most current work of our laboratory and others and describes many of the laboratory findings behind our current understanding of the PN enteral environment. Understanding the interaction between nutrient delivery, the intestinal microbiome, and PN-associated complications may lead to the development of novel therapies to enhance safety and quality of life for patients requiring PN.

Altered fecal short chain fatty acid composition in children with a history of Hirschsprung-associated enterocolitis

PURPOSE Children with Hirschsprung disease (HD) who have a history of enterocolitis (HAEC) have a shift in colonic microbiota, many of which are necessary for short chain fatty acid (SCFA) production. As SCFAs play a critical role in colonic mucosal preservation, we hypothesized that fecal SCFA composition is altered in children with HAEC. METHODS A multicenter study enrolled 18 HD children, abstracting for history of feeding, antibiotic/probiotic use, and enterocolitis symptoms. HAEC status was determined per Pastor et al. criteria (12). Fresh feces were collected for microbial community analysis via 16S sequencing as well as SCFA analysis by gas chromatography-mass spectrometry. RESULTS Nine patients had a history of HAEC, and nine had never had HAEC. Fecal samples from HAEC children showed a 4-fold decline in total SCFA concentration vs. non-HAEC HD patients. We then compared the relative composition of individual SCFAs and found reduced acetate and increased butyrate in HAEC children. Finally, we measured relative abundance of SCFA-producing fecal microbiota. Interestingly, 10 of 12 butyrate-producing genera as well as 3 of 4 acetate-producing genera demonstrated multi-fold expansion. CONCLUSION Children with HAEC history have reduced fecal SCFAs and altered SCFA profile. These findings suggest a complex interplay between the colonic metabolome and changes in microbiota, which may influence the pathogenesis of HAEC.

TPN-associated intestinal epithelial cell atrophy is modulated by TLR4/EGF signaling pathways

Recent studies suggest a close interaction between epidermal growth factor (EGF) and TLR signaling in the modulation of intestinal epithelial cell (IEC) proliferation; however, how these signaling pathways adjust IEC proliferation is poorly understood. We utilized a model of total parenteral nutrition (TPN), or enteral nutrient deprivation, to study this interaction as TPN results in mucosal atrophy due to decreased IEC proliferation and increased apoptosis. We identified the novel finding of decreased mucosal atrophy in TLR4 knockout (TLR4KO) mice receiving TPN. We hypothesized that EGF signaling is preserved in TLR4KO‐TPN mice and prevents mucosal atrophy. C57B1/6 and strain‐matched TLR4KO mice were provided either enteral feeding or TPN. IEC proliferation and apoptosis were measured. Cytokine and growth factor abundances were detected in both groups. To examine interdependence of these pathways, ErbB1 pharmacologic blockade was used. The marked decline in IEC proliferation with TPN was nearly prevented in TLR4KO mice, and intestinal length was partially preserved. EGF was significantly increased, and TNF‐α decreased in TLR4KO‐TPN versus wild‐type (WT)‐TPN mice. Apoptotic positive crypt cells were 15‐fold higher in WT‐TPN versus TLR4KO‐TPN mice. Bcl‐2 was significantly increased in TLR4KOTPN mice, while Bax decreased 10‐fold. ErbB1 blockade prevented this otherwise protective effect in TLR4KO‐sTPN mice. TLR4 blockade significantly prevented TPN‐associated atrophy by preserving proliferation and preventing apoptosis. This is driven by a reduction in TNF‐α abundance and increased EGF. Potential manipulation of this regulatory pathway may have significant clinical potential to prevent TPN‐associated atrophy.—Freeman, J. J., Feng, Y., Demehri, F. R., Dempsey, P. J., Teitelbaum, D. H. TPN‐associated intestinal epithelial cell atrophy is modulated by TLR4/EGF signaling pathways. FASEB J. 29, 2943‐2958 (2015). www.fasebj.org

Small Bowel Diameter in Short Bowel Syndrome as a Predictive Factor for Achieving Enteral Autonomy

Children with short bowel syndrome commonly have dilated small bowel. We found that the extent of dilation was associated with bowel length and that both were related to achieving enteral autonomy.

Development of an endoluminal intestinal attachment for a clinically applicable distraction enterogenesis device.

PURPOSE Previous methods of distraction enterogenesis have relied upon blind-ending intestinal segments or transmural device fixation, requiring multiple operations and potential bowel injury. We hypothesized that using a novel attachment would allow reversible device coupling to the luminal bowel surface, achieving effective endoluminal distraction. METHODS A telescopic hydraulic device was designed with latex balloon attachments covered with high-friction mesh and a dilating fenestrated elastic mask (DFM attachment), allowing mesh-to-mucosa contact only with inflation. Yorkshire pigs underwent jejunal Roux-en-Y limb creation and device placement via jejunostomy. Devices underwent 3 cycles of balloon inflation and hydraulic extension/retraction per day for 7 days and then explanted and studied for efficacy. RESULTS DFM attachment allowed reversible, high-strength endoluminal coupling without tissue injury or reduction in bowel perfusion. After 7 day implant, distracted bowel achieved a 44 ± 2% increase in length vs. fed, nondistracted bowel, corresponding to a gain of 7.1 ± 0.3 cm. Distracted bowel demonstrated increased epithelial cell proliferation vs. control bowel. Attachment sites demonstrated villus flattening, increased crypt depth, thicker muscularis mucosa, and unchanged muscularis propria thickness vs. CONCLUSION Novel high-strength, reversible attachments enabled fully endoluminal distraction enterogenesis, achieving length gains comparable to open surgical techniques. This approach may allow development of clinically applicable technology for SBS treatment.

Analysis of risk factors contributing to morbidity from gastrojejunostomy feeding tubes in children

PURPOSE The purpose of this study was to define morbidity from gastrojejunostomy tube (GJT) placement in children. METHODS A retrospective single-center 5-year review of GJT placement in children was performed. Age, weight, prior surgery, indication, type of GJT, and complications (GJT replacement, wound complications, and perforation) were recorded. Logistic regression for morbidity was performed. RESULTS 142 children underwent 394 GJT placements at a median age of 2.7years (range 5 weeks-18years). The most common indications were failure to thrive (62%) and reflux (25%). Among the 296 GJT replacements, the most common reason was tube dislodgement (30%). Risk factors for replacement, which occurred at a median interval of 12 weeks (range 2days-2.4years), were peristomal complaint (OR=5.4, p=0.02) and prior GJT replacement (OR=1.8, p=0.03). In all, 7 (5%) jejunal perforations occurred at a median of 3 days (range 0-21 days) from GJT placement. Patients with perforation had a median weight of 4.6kg (range 3-11.2kg) and age of 3.9months (range 8 weeks-2.1years). Lower weight (p<0.01) and younger age (p=0.02) predicted perforation, with those weighing less than 6kg (OR=51.9, p<0.001) or younger than 6months (OR=28.6, p<0.01) at highest risk. CONCLUSIONS GJT placement has a significant risk of recurrent dislodgement and the highest risk of perforation in children weighing less than 6kg or younger than 6months. Alternate feeding options should be strongly considered in this vulnerable population.