Yasuhiro Torashima

Human tissue-engineered small intestine forms from postnatal progenitor cells.

PURPOSE Tissue-engineered small intestine (TESI) represents a potential cure for short bowel syndrome (SBS). We previously reported full-thickness intestine formation using an organoid units-on-scaffold approach in rodent and swine models. Transplanted intestinal xenografts have been documented to survive from human fetal tissue but not from postnatal tissue. We now present the first report of human TESI from postnatal tissue. METHODS Organoid units (OU) were prepared from human small bowel resection specimens, loaded onto biodegradable scaffolds and implanted into NOD/SCID gamma chain-deficient mice. After 4 weeks, TESI was harvested and immunostained for β2-microglobulin to identify human tissue, villin for enterocytes, lysozyme for Paneth cells, chromogranin-A for enteroendocrine cells, mucin-2 for goblet cells, smooth muscle actin and desmin to demonstrate muscularis, and S-100 for nerves. RESULTS All TESI was of human origin. Immunofluorescence staining of human TESI reveals the presence of all four differentiated cell types of mature human small intestine, in addition to the muscularis and the supporting intestinal subepithelial myofibroblasts. Nerve tissue is also present. CONCLUSIONS Our technique demonstrates survival, growth, and differentiation of postnatally derived human small intestinal OU into full thickness TESI in murine hosts. This regenerative medicine strategy may eventually assist in the treatment of SBS.

The risk factors and predictive factors for anastomotic leakage after resection for colorectal cancer: reappraisal of the literature

Anastomotic leakage is a serious complication that can occur after colorectal surgery. Several risk factors for anastomotic leakage have been reported based on the findings of prospective and retrospective studies, including patient characteristics, the use of neoadjuvant therapy, the tumor location, intraoperative events, etc. However, as these risk factors affect each other, the statistical results have differed in each study. In addition, differences in surgical methods, including laparoscopy versus laparotomy or stapling anastomosis versus handsewn anastomosis, may influence the incidence of anastomotic leakage. This mini-review summarizes the results of reported papers to clarify the current evidence of risk factors for anastomotic leakage.

Tissue engineering of the intestine in a murine model.

Tissue-engineered small intestine (TESI) has successfully been used to rescue Lewis rats after massive small bowel resection, resulting in return to preoperative weights within 40 days.(1) In humans, massive small bowel resection can result in short bowel syndrome, a functional malabsorptive state that confers significant morbidity, mortality, and healthcare costs including parenteral nutrition dependence, liver failure and cirrhosis, and the need for multivisceral organ transplantation.(2) In this paper, we describe and document our protocol for creating tissue-engineered intestine in a mouse model with a multicellular organoid units-on-scaffold approach. Organoid units are multicellular aggregates derived from the intestine that contain both mucosal and mesenchymal elements,(3) the relationship between which preserves the intestinal stem cell niche.(4) In ongoing and future research, the transition of our technique into the mouse will allow for investigation of the processes involved during TESI formation by utilizing the transgenic tools available in this species.(5)The availability of immunocompromised mouse strains will also permit us to apply the technique to human intestinal tissue and optimize the formation of human TESI as a mouse xenograft before its transition into humans. Our method employs good manufacturing practice (GMP) reagents and materials that have already been approved for use in human patients, and therefore offers a significant advantage over approaches that rely upon decellularized animal tissues. The ultimate goal of this method is its translation to humans as a regenerative medicine therapeutic strategy for short bowel syndrome.

Human tissue-engineered colon forms from postnatal progenitor cells: an in vivo murine model

AIM Loss of colon reservoir function after colectomy can adversely affect patient outcomes. In previous work, human fetal intestinal cells developed epithelium without mesenchyme following implantation in mice. However, for humans, postnatal tissue would be the preferred donor source. We generated tissue-engineered colon (TEC) from postnatal human organoid units. MATERIALS & METHODS Organoid units were prepared from human colon waste specimens, loaded onto biodegradable scaffolds and implanted into immunocompromised mice. After 4 weeks, human TEC was harvested. Immunofluorescence staining confirmed human origin, identified differentiated epithelial cell types and verified the presence of supporting mesenchyme. RESULTS Human TEC demonstrated a simple columnar epithelium. Immunofluorescence staining demonstrated human origin and the three differentiated cell types of mature colon epithelium. Key mesenchymal components (smooth muscle, intestinal subepithelial myofibroblasts and ganglion cells) were seen. CONCLUSION Colon can form from human progenitor cells on a scaffold in a mouse host. This proof-of-concept experiment is an important step in transitioning TEC to human therapy.

A “Living Bioreactor” for the Production of Tissue-Engineered Small Intestine

Here, we describe the use of a mouse model as a living bioreactor for the generation of tissue-engineered small intestine. Small intestine is harvested from donor mice with subsequent isolation of organoid units (a cluster of mesenchymal and epithelial cells). Some of these organoid units contain pluripotent stem cells with a preserved relationship with the mesenchymal stem cell niche. A preparation of organoid units is seeded onto a biodegradable scaffold and implanted intraperitoneally within the omentum of the host animal. The cells are nourished initially via imbibition until neovascularization occurs. This technique allows the growth of fully differentiated epithelium (composed of Paneth cells, goblet cells, enterocytes and enteroendocrine cells), muscle, nerve, and blood vessels of donor origin. Variations of this technique have been used to generate tissue-engineered stomach, large intestine, and esophagus. The variations include harvest technique, length of digestion, and harvest times.

Rectal Cancer with Paraneoplastic Nephropathy: Association of Vascular Endothelial Growth Factor

A patient with advanced rectal cancer was complicated by progressing proteinuria and hypoproteinemia. Low anterior resection was a procedure of choice. A surgical specimen obtained by intraoperative renal biopsy showed the findings of minimal change nephrotic syndrome. After surgery, nephropathy remitted promptly and completely. Her pre/postoperative serum level of vascular endothelial growth factor was 1,880/52.3 pg/ml, suggesting its elevation was associated with the nephropathy. Immunohistochemistry revealed strongly expressed tumor vascular endothelial cell growth factor. Minimal change nephrotic syndrome is a rare type of paraneoplastic nephropathy, and successful remission may require therapeutic resection of the underlying tumor, or administration of a vascular endothelial growth factor antagonist if the tumor is unresectable.

Surgery for ileal mesenteric lymphangioma during pregnancy: Case report and review of the literature

Mesenteric lymphangioma is one of the least frequently encountered types of benign tumor. This case report concerns a 31-year-old pregnant woman with a mesenteric cystic lymphangioma in the ileum. The multiloculated cystic mass was noted near the uterus by CT before the patient became pregnant. After becoming pregnant, she was followed without treatment for the asymptomatic mass. At 25 weeks’ gestation, however, she underwent emergency surgical treatment for small bowel obstruction. Concomitant small bowel resection was performed to remove the cyst. Herein we review seven reported cases of mesenteric benign tumor in pregnancy and explore the clinical features.

Fgf10 overexpression enhances the formation of tissue-engineered small intestine.

Short bowel syndrome (SBS) is a morbid and mortal condition characterized in most patients by insufficient intestinal surface area. Current management strategies are inadequate, but tissue‐engineered small intestine (TESI) offers a potential therapy. A barrier to translation of TESI is the generation of scalable mucosal surface area to significantly increase nutritional absorption. Fibroblast growth factor 10 (Fgf10) is a critical growth factor essential for the development of the gastrointestinal tract. We hypothesized that overexpression of Fgf10 would improve the generation of TESI. Organoid units, the multicellular donor tissue that forms TESI, were derived from Rosa26rtTA/+, tet(o)Fgf10/– or Fgf10Mlc‐nlacZ‐v24 (hereafter called Fgf10lacZ ) mice. These were implanted into the omentum of NOD/SCID γ‐chain‐deficient mice and induced with doxycycline in the case of tet(o)Fgf10/–. Resulting TESI were explanted at 4 weeks and studied by histology, quantitative RT–PCR and immunofluorescence. Four weeks after implantation, Fgf10 overexpressing TESI was larger and weighed more than the control tissues. Within the mucosa, the villus height was significantly longer and crypts contained a greater percentage of proliferating epithelial cells. A fully differentiated intestinal epithelium with enterocytes, goblet cells, enteroendocrine cells and Paneth cells was identified in the Fgf10‐overexpressing TESI, comparable to native small intestine. β‐Galactosidase expression was found in both the epithelium and the mesenchyme of the TESI derived from the Fgf10LacZ duodenum. However, this was not the case with TESI generated from jejunum and ileum. We conclude that Fgf10 enhances the formation of TESI. Copyright

Suppression of reactive oxygen species develops lymph node metastasis in colorectal cancer.

BACKGROUND/AIMS Recent evidence indicates that reactive oxygen species (ROS) can induce a wide type of cellular responses from proliferation to senescence and cell death. ROS may not be an absolute carcinogenic factor or cancer suppressor. The aim of this study was to assess the biological paradox of ROS in colorectal cancer cells. METHODOLOGY Blood specimens were obtained from the drainage vein of the tumor during operation in 135 patients with colorectal cancer. Serum ROS levels were measured using the derivatives of reactive oxygen metabolites (d-ROM) test. RESULTS Serum ROS levels increased significantly in tumor size larger than 40mm (p<0.01). On the other hand, serum ROS levels decreased significantly in patients with lymph node metastasis (p<0.01). Multiple linear regression models showed a significant association of serum ROS levels with serum carcinoembryonic antigen (CEA) levels (p<0.01) and lymph node metastasis (p=0.026). CONCLUSIONS In colorectal cancer cells, the increase of intracellular ROS is first associated with cell growth and invasion. However, a further increase inhibits cancer cell proliferation, whereas any decrease in ROS concentration needs to stimulate lymph node metastasis. Thus, a precise understanding how ROS are generated and involved in lymph node metastasis will help us to design better therapeutic strategies.

Prediction and management of a low-lying costal arch which restricts the operative working space during laparoscopic cholecystectomy.

Background/purposeLaparoscopic cholecystectomy is difficult to perform in patients with a low-lying costal arch that entirely covers the liver. We conducted this study to clarify the factors related to a low-lying costal arch and establish countermeasures to circumvent this characteristic.MethodsThe study included 103 consecutive patients who underwent a laparoscopic cholecystectomy. The possible clinical factors associated with a low-lying costal arch restricting the operative working space were analyzed. The position of the liver against the costal arch and the presumed surgical visual angle for laparoscopic cholecystectomy, comprising the hepatic porta, umbilicus, and costal arch, were estimated with abdominal multidetector computed tomography (MDCT).ResultsSeven (7%) patients had a low-lying costal arch presenting an inadequate exposure of Calot’s triangle and restricted instrument mobility during laparoscopic cholecystectomy, and three patients required conversion to a laparotomy. A low-lying costal arch was significantly associated with advanced age, shorter stature, lighter body weight, coexisting kyphoscoliosis, gallbladder pathology, laparotomy conversion, and most of all, the liver edge lying above the costal arch and a narrow surgical visual angle upon MDCT. Of the seven patients with a critical low-lying costal arch, four underwent a successful laparoscopic cholecystectomy, this being done by lifting the right costal arch to create a workable surgical field; the rib-lifting procedure was planned as part of the scheduled procedure in the other three patients because the preoperative MDCT examination indicated a poor working space for a laparoscopic cholecystectomy.ConclusionsA low-lying costal arch is a substantial risk factor for conversion to a laparotomy when performing a laparoscopic cholecystectomy. However, the operative difficulty related to a low-lying costal arch can be predicted by using preoperative MDCT images and can be managed with proper planning and the appropriate use of the rib-lifting technique.