Rotavirus Reassortant-Induced Murine Model of Liver Fibrosis Parallels Human Biliary Atresia.

BACKGROUND AND AIMS: Biliary atresia (BA) is a devastating neonatal cholangiopathy that progresses to fibrosis and end-stage liver disease by 2 years of age. Portoenterostomy may reestablish biliary drainage, but, despite drainage, virtually all afflicted patients develop fibrosis and progress to end-stage liver disease requiring liver transplantation for survival. APPROACH AND RESULTS: In the murine model of BA, rhesus rotavirus (RRV) infection of newborn pups results in a cholangiopathy paralleling human BA and has been used to study mechanistic aspects of the disease. Unfortunately, nearly all RRV-infected pups succumb by day of life 14. Thus, in this study we generated an RRV-TUCH rotavirus reassortant (designated as TR(VP2,VP4) ) that when injected into newborn mice causes an obstructive jaundice phenotype with lower mortality rates. Of the mice that survived, 63% developed Ishak stage 3-5 fibrosis with histopathological signs of inflammation/fibrosis and bile duct obstruction. CONCLUSIONS: This model of rotavirus-induced neonatal fibrosis will provide an opportunity to study disease pathogenesis and has potential to be used in preclinical studies with an objective to identify therapeutic targets that may alter the course of BA.

The SRL peptide of rhesus rotavirus VP4 protein governs cholangiocyte infection and the murine model of biliary atresia.

Biliary atresia (BA) is a neonatal obstructive cholangiopathy that progresses to end-stage liver disease, often requiring transplantation. The murine model of BA, employing rhesus rotavirus (RRV), parallels human disease and has been used to elucidate mechanistic aspects of a virus induced biliary cholangiopathy. We previously reported that the RRV VP4 gene plays an integral role in activating the immune system and induction of BA. Using rotavirus binding and blocking assays, this study elucidated how RRV VP4 protein governs cholangiocyte susceptibility to infection both in vitro and in vivo in the murine model of BA. We identified the amino acid sequence on VP4 and its cholangiocyte binding protein, finding that the sequence is specific to those rotavirus strains that cause obstructive cholangiopathy. Pretreatment of murine and human cholangiocytes with this VP4-derived peptide (TRTRVSRLY) significantly reduced the ability of RRV to bind and infect cells. However, the peptide did not block cholangiocyte binding of TUCH and Ro1845, strains that do not induce murine BA. The SRL sequence within TRTRVSRLY is required for cholangiocyte binding and viral replication. The cholangiocyte membrane protein bound by SRL was found to be Hsc70. Inhibition of Hsc70 by small interfering RNAs reduced RRV's ability to infect cholangiocytes. This virus-cholangiocyte interaction is also seen in vivo in the murine model of BA, where inoculation of mice with TRTRVSRLY peptide significantly reduced symptoms and mortality in RRV-injected mice. CONCLUSION: The tripeptide SRL on RRV VP4 binds to the cholangiocyte membrane protein Hsc70, defining a novel binding site governing VP4 attachment. Investigations are underway to determine the cellular response to this interaction to understand how it contributes to the pathogenesis of BA. (Hepatology 2017;65:1278-1292).

Rhesus rotavirus VP4 sequence-specific activation of mononuclear cells is associated with cholangiopathy in murine biliary atresia.

Biliary atresia (BA), a neonatal obstructive cholangiopathy, remains the most common indication for pediatric liver transplantation in the United States. In the murine model of BA, Rhesus rotavirus (RRV) VP4 surface protein determines biliary duct tropism. In this study, we investigated how VP4 governs induction of murine BA. Newborn mice were injected with 16 strains of rotavirus and observed for clinical symptoms of BA and mortality. Cholangiograms were performed to confirm bile duct obstruction. Livers and bile ducts were harvested 7 days postinfection for virus titers and histology. Flow cytometry assessed mononuclear cell activation in harvested cell populations from the liver. Cytotoxic NK cell activity was determined by the ability of NK cells to kill noninfected cholangiocytes. Of the 16 strains investigated, the 6 with the highest homology to the RRV VP4 (>87%) were capable of infecting bile ducts in vivo. Although the strain Ro1845 replicated to a titer similar to RRV in vivo, it caused no symptoms or mortality. A Ro1845 reassortant containing the RRV VP4 induced all BA symptoms, with a mortality rate of 89%. Flow cytometry revealed that NK cell activation was significantly increased in the disease-inducing strains and these NK cells demonstrated a significantly higher percentage of cytotoxicity against noninfected cholangiocytes. Rotavirus strains with >87% homology to RRV's VP4 were capable of infecting murine bile ducts in vivo. Development of murine BA was mediated by RRV VP4-specific activation of mononuclear cells, independent of viral titers.

Laparoscopic donor nephrectomy for the pediatric recipient population: Risk factors for adverse outcomes.

Kidney transplantation is the optimal treatment of ESRD in children. Some studies have reported inferior outcomes in recipients of LDN allografts who are ≤ 5 yr of age. We performed a retrospective review of pediatric recipient outcomes of 110 LDN allografts at our institution and examined predictors of adverse outcomes. Subgroup analysis was performed by dividing recipients into three age categories: 0-5 yr, 6-17 yr, and ≥ 18 yr. There was no significant difference between incidences of DGF or ARE between groups. Kaplan-Meier analysis demonstrated 100% allograft survival in 0- to 5-yr-old recipients, nearly reaching statistical significance (p = 0.07) for outcome superior to that of the two older age groups. Pretransplant HD was associated with increased risk of DGF (p = 0.05). Significant risk factors for ARE were recipient weight >15 kg (p = 0.033) and multiple renal arteries (p = 0.047). Previous ARE was associated with an increased risk of allograft failure (p = 0.02). LDN is not associated with increased risk of DGF, ARE, or allograft failure in the youngest recipients. These findings support an aggressive pursuit of preemptive transplantation even in the youngest pediatric allograft recipients.

Multimodal therapy including liver transplantation for hepatic undifferentiated embryonal sarcoma.

The outcomes of hepatic undifferentiated embryonal sarcoma (HUES) have historically been limited by persistent, unresectable disease and the subsequent development of disease resistance and dissemination. We present our institutional experience with HUES and assess current treatment trends and outcomes in the era of liver transplantation. We conducted a retrospective chart review of cases presenting with HUES at our institution over the past 10 years. The collected data included age, sex, presenting symptoms, imaging and the associated Pretreatment Extent of Disease (PRETEXT) score, pathology, chemotherapy, surgical interventions, and outcomes. Approval was obtained from the institutional review board of the Cincinnati Children's Hospital Medical Center. HUES was identified in 6 patients (4 males and 2 females) with a median age at diagnosis of 11 years (range = 7-13 years). Initial imaging was available for all but 1 patient. The PRETEXT stage for these patients ranged from II to III. One patient was diagnosed with lung metastases. Two patients underwent upfront resection, and 1 patient received neoadjuvant therapy and then conventional resection. Three patients were treated with orthotopic liver transplantation (OLT) after neoadjuvant chemotherapy (primary OLT in 2 cases and salvage OLT for local recurrence in 1 case). Two patients received posttransplant adjuvant chemotherapy. All 6 patients remained in clinical remission with a mean follow-up of 35 months (range = 12-84 months). In conclusion, OLT has rarely been reported as a treatment option for HUES. The addition of liver transplantation as a surgical option for treating patients with HUES can result in improved survival for patients whose tumors are initially unresectable or recur.

Physiology of the small intestine after resection and transplant.

PURPOSE OF REVIEW: Recent studies have evaluated intestinal physiology following bowel resection; understanding changes in small bowel physiology after intestinal transplantation has received less attention. In this review, we will examine recent studies focused on changes in intestinal physiology following resection and intestinal transplantation. RECENT FINDINGS: Absorption, immunity, and motility are fundamental components of small bowel physiology. Absorption after resection or transplantation is mediated by adaptation and enterocyte function. After resection, adaptation results in increased villus height and crypt depth. Hepatocyte growth factor and epidermal growth factors cause enterocyte hypertrophy and hyperplasia, allowing greater peptide uptake. Little is known about intestinal adaptation after transplant, but enteral autonomy is attainable. Immunity in small bowel after transplantation relies on a balance of innate and adaptive immune responses in the presence of the luminal microbiota. Intraepithelial lymphocytes are decreased following small bowel resection. After small bowel transplant, the number and the ratio of intraepithelial lymphocytes to enterocytes, as well as changes in the microbiota, can be used to identify rejection. After intestinal transplant, immune-mediated dysmotility is common. Perioperative infliximab in addition to tacrolimus may decrease the inflammation that contributes to dysmotility. SUMMARY: As intestinal transplantation becomes more successful, understanding how absorption, immunity, and motility changes will allow for optimization of bowel function.

Role of myeloid differentiation factor 88 in Rhesus rotavirus-induced biliary atresia.

BACKGROUND: Biliary atresia (BA) is a unique neonatal disease resulting from inflammatory and fibrosing obstruction of the extrahepatic biliary tree. Previous studies have demonstrated the critical role of innate immunity and the Th1 response to activated inflammatory cells and overexpressed cytokines in the pathogenesis of BA. Myeloid differentiation factor 88 (MyD88) is a critical adaptor molecule that has been shown to play a crucial role in immunity. We investigated the role of MyD88 in the inflammatory response and development of cholangiopathy in murine BA. METHODS: MyD88 knockout (MyD88(-/-)) and wild-type (WT) BALB/c pups were injected with Rhesus rotavirus or saline on day 1 of life. The mice were monitored for clinical symptoms of BA, including jaundice, acholic stools, bilirubinuria, and death. The liver and extrahepatic bile ducts were harvested for histologic evaluation and the quantification of viral content, determination of cytokine expression, and detection of inflammatory cells. RESULTS: Rhesus rotavirus infection produced symptoms in 100% of both MyD88(-/-) and WT pups, with survival of 18% of WT and 0% of MyD88(-/-) mice. Histologic analysis demonstrated bile duct obstruction in both MyD88(-/-) and WT mice. Viral titers obtained 7 d after infection and expression of interferon-γ and tumor necrosis factor-α at day 3, 5, 8, and 12 after infection revealed no significant differences between the WT and MyD88(-/-) mice. Flow cytometry demonstrated similar levels of activated CD8+ T cells and natural killer cells. CONCLUSIONS: The pathogenesis of murine BA is independent of the MyD88 signaling inflammatory pathway, suggesting alternative mechanisms are crucial in the induction of the model.

Large Traumatic Lumbar Hernia: A Difficult and Uncommon Problem.

Lumbar hernias are congenital or acquired posterolateral abdominal wall hernias and are located in the superior or inferior lumbar triangle. Traumatic lumbar hernias are rare, and the optimal method to repair these is not well-defined. We present the case of a 59-year-old obese female who presented after a motor vehicle collision with an 8.8 cm traumatic right-sided inferior lumbar hernia and overlying complex abdominal wall laceration. The patient underwent an open repair with retro rectus polypropylene mesh and biologic mesh underlay several months after the abdominal wall wound healed, and the patient lost 60 pounds. The patient recovered well without complications or recurrence at the one-year follow-up. This case demonstrates a complex, open surgical approach to repair a large traumatic lumbar hernia not amenable to laparoscopic repair.

Rotavirus infection of human cholangiocytes parallels the murine model of biliary atresia.

INTRODUCTION: Biliary atresia (BA) is the leading indication for liver transplantation in the pediatric population. The murine model of BA supports a viral etiology, because infection of neonatal mice with rhesus rotavirus (RRV) results in biliary obstruction. Viral infection targets the biliary epithelium and development of the model is viral strain dependent. No study has yet determined whether human cholangiocytes are also susceptible to rotaviral infection. We established an in vitro human model using an immortalized human cholangiocyte cell line and primary human cholangiocytes obtained from explanted livers to determine human cholangiocyte susceptibility to rotavirus infection. METHODS: Replication and binding assays were performed on immortalized mouse (mCL) and human (H69) cells using six different strains of rotavirus. Primary human cholangiocytes were isolated from cadaveric livers, characterized in culture, and infected with RRV, which causes BA in mice, and another simian strain, TUCH, which does not cause BA in mice. RESULTS: Immortalized mouse and human cholangiocytes demonstrated similar patterns of infectivity and binding with different strains of rotavirus. Both cell lines produced a significantly higher viral yield with RRV infection than with the other strains tested. In primary human cholangiocytes, which maintained their epithelial characteristics, as demonstrated by cytokeratin staining, RRV replicated to a yield 1000-fold higher than TUCH. CONCLUSIONS: Both immortalized and primary human cholangiocytes are susceptible to RRV infection in a fashion similar to murine cholangiocytes. These novel findings suggest rotavirus infection could have a potential role in the pathogenesis of human BA.

Large Retroperitoneal Hematoma: A Rare Intraoperative Complication of Total Vaginal Hysterectomy.

Retroperitoneal (RP) hematoma is a rare complication of total vaginal hysterectomy. A 45-year-old female G4P3013 with a history of abnormal uterine bleeding refractory to treatment by endometrial ablation and stress urinary incontinence underwent total vaginal hysterectomy, bilateral salpingectomy, bilateral uterosacral ligament suspension, anterior colporrhaphy, and cystoscopy. After the hysterectomy the left uterine artery pedicle was hemostatic; however, the patient became hemodynamically unstable and anemic. Laparoscopy revealed a stable zone III RP hematoma. Intraoperative observation revealed no further expansion of the hematoma. Left iliac angiography and aortography revealed there was no extravasation from the uterine arteries and gonadal vessels. Four days post-operative abdominal CT showed a stable hematoma. Hemodynamic instability resolved over the post-operative course. RP hematoma must be included in the differential for the evaluation of acute intraoperative hemodynamic instability with an unclear source.

Effect of a 0.5-T static magnetic field on conduction in guinea pig spinal cord.

Compound-evoked potentials were recorded from excised adult guinea pig spinal cords before, during, and following exposure to a 0.5-T static magnetic field (SMF). There was no change in response latency during exposure but there was a small, statistically significant, decrease in amplitude. Maximum effect was evident 1 to 2 min after the field was turned on with return to baseline within 1 min after the field was turned off. These results may be explained by a conduction block in the small axon subpopulation due to the effect of static magnetic fields on voltage-activated sodium channels. The relative selectivity of the field is believed to occur because of the relatively greater number of sodium channels present in smaller axons.

Rotavirus replication in the cholangiocyte mediates the temporal dependence of murine biliary atresia.

Biliary atresia (BA) is a neonatal disease that results in obliteration of the biliary tree. The murine model of BA, which mirrors the human disease, is based upon infection of newborn mice with rhesus rotavirus (RRV), leading to an obstructive cholangiopathy. The purpose of this study was to characterize the temporal relationship between viral infection and the induction of this model. BALB/c mice were infected with RRV on day of life (DOL) 0, 3, 5, and 7. Groups were characterized as early-infection (infection by DOL 3) or late-infection (infection after DOL 5). Early RRV infection induced symptoms in 95% of pups with a mortality rate of 80%. In contrast, late infection caused symptoms in only 50% of mice, and 100% of pups survived. The clinical findings correlated with histological analysis of extrahepatic biliary trees, cytokine expression, and viral titers. Primary murine cholangiocytes isolated, cultured, and infected with RRV yielded higher titers of infectious virus in those harvested from DOL 2 versus DOL 9 mice. Less interferon alpha and beta was produced in DOL 2 versus DOL 9 RRV infected primary cholangiocytes. Injection of BALB/c interferon alpha/beta receptor knockout (IFN-αßR(-/-)) pups at DOL 7 showed increased symptoms (79%) and mortality (46%) when compared to late infected wild type mice. In conclusion, the degree of injury sustained by relatively immature cholangiocytes due to more robust RRV replication correlated with more severe clinical manifestations of cholangiopathy and higher mortality. Interferon alpha production by cholangiocytes appears to play a regulatory role. These findings confirm a temporal dependence of RRV infection in murine BA and begin to define a pathophysiologic role of the maturing cholangiocyte.