Shawn D. Safford

Neurogenic differentiation of murine and human adipose-derived stromal cells

The identification of cells capable of neuronal differentiation has great potential for cellular therapies. We examined whether murine and human adipose-derived adult stem (ADAS) cells can be induced to undergo neuronal differentiation. We isolated ADAS cells from the adipose tissue of adult BalbC mice or from human liposuction tissue and induced neuronal differentiation with valproic acid, butylated hydroxyanisole, insulin, and hydrocortisone. As early as 1-3 h after neuronal induction, the phenotype of ADAS cells changed towards neuronal morphology. Following neuronal induction, muADAS cells displayed immunocytochemical staining for GFAP, nestin and NeuN and huADAS cells displayed staining for intermediate filament M, nestin, and NeuN. Following neuronal induction of murine and human ADAS cells, Western blot analysis confirmed GFAP, nestin, and NeuN protein expression. Pretreatment with EGF and basic FGF augmented the neuronal differentiation of huADAS cells. The neuronal differentiation of stromal cells from adipose tissue has broad biological and clinical implications.

Characterization of neuronal/glial differentiation of murine adipose-derived adult stromal cells

Neural tissue has limited capacity for intrinsic repair after injury, and the identification of alternate sources of neuronal stem cells has broad clinical potential. Preliminary studies have demonstrated that adipose-derived adult stromal (ADAS) cells are capable of differentiating into mesenchymal and non-mesenchymal cells in vitro, including cells with select characteristics of neuronal/glial tissue. In this study, we extended these observations to test the hypothesis that murine (mu) ADAS cells can be induced to exhibit characteristics of neuronal and glial tissue by exposure to a cocktail of induction agents. We characterized the differentiation of muADAS cells in vitro using immunohistochemistry and immunoblotting, and examined whether these cells respond to the glutamate agonist N-methyl-D-aspartate (NMDA). We found that induced muADAS cells express proteins indicative of neuronal/glial cells, including nestin, GFAP, S-100, NeuN, MAP2, tau, and beta-III tubulin. Induced muADAS cells express gamma-aminobutyric acid (GABA), the NR-1 and NR-2 subunits of the glutamate receptor, GAP-43, synapsin I, and voltage-gated calcium channels. Finally, induced muADAS cells demonstrate decreased viability in response to NMDA. These findings suggest that muADAS cells can be induced to exhibit several phenotypic, morphologic, and excitotoxic characteristics consistent with developing neuronal and glial tissue.

Iodine -131 metaiodobenzylguanidine is an effective treatment for malignant pheochromocytoma and paraganglioma☆

INTRODUCTION Iodine 131-meta-iodobenzylguanidine ((131)I-MIBG) has been applied to the palliative treatment of metastatic pheochromocytoma in small studies. We report our institutional experience for the treatment of metastatic pheochromocytoma and paraganglioma. METHODS We performed a retrospective review of 33 patients with metastatic pheochromocytoma (n=22) and paraganglioma (n=11) treated at our institution with (131)I-MIBG over a 10-year period. RESULTS Patients received a mean dose of 388+/-131 mCi (131)I-MIBG. Median survival after treatment was 4.7 years. Most patients experienced a symptomatic response leading to an improved survival (4.7 years vs 1.8 years, P<.01). Patients with a measurable hormone response demonstrated an increased survival in comparison to those with no response (4.7 years vs 2.6 years, P=.01). Patients who received a high dose (>500 mCi) as their initial therapy also had improved survival (3.8 years vs 2.8 years, P=.02). CONCLUSION These data support (131)I-MIBG treatment for select patients with metastatic pheochromocytoma. In our experience, prolonged survival was best predicted by symptomatic and hormone response to (131)I-MIBG treatment. An initial dose of 500 mCi may be optimal. The benefit of (131)I-MIBG treatment for metastatic pheochromocytoma must also be weighed against its side effects.

High Hospital Volume Is Associated with Better Outcomes for Breast Cancer Surgery: Analysis of 233,247 Patients

Background:The relationship between hospital volume and outcomes needs to be further elucidated for low-risk procedures such as surgical therapy of localized breast cancer. The objective of this investigation was to assess the relationship between hospital volume and outcomes for breast cancer surgery.Methods: A total of 233,247 patients who underwent breast-conserving therapy (BCT) and breast-ablative therapy (BAT) for localized breast cancer were extracted from 13 years (1988–2000) of the Nationwide Inpatient Samples. Hospital volume was classified as low (<30 cases/year), intermediate (≥ 30 to <70cases/year), and high (≥ 70 cases/year). Multiple linear and logistic regression analyses were used to assess the risk-adjusted association between hospital volume and outcomes.Results: In risk-adjusted analyses, patients operated on at low-volume hospitals were 3.04 (p = 0.03) times more likely to die after BCT compared with patients operated on at high-volume hospitals. Similarly, low-volume hospitals had a significantly higher likelihood of postoperative complications (odds ratio [OR] = 1.73, p = 0.01 for BCT; OR = 1.44, p < 0.001 for BAT) compared with high-volume hospitals. Compared with low-volume hospitals, length of hospital stay was significantly shorter and nonroutine patient discharge significantly lower for high-volume providers for both BCT and BAT (all p < 0.001). Patients were also significantly less likely to undergo BCT if operated on in a low- or intermediate-volume hospital compared with a high-volume provider (p < 0.001).Conclusions: High-volume hospitals had significantly lower nonroutine patient discharge, postoperative morbidity and mortality, shorter length of hospital stay, and higher likelihood of performing BCT. Referral of patients with localized breast cancer to high-volume hospitals may be justified.

Longitudinal mechanical tension induces growth in the small bowel of juvenile rats

Introduction: The aim of our study was to apply longitudinal force to the small bowel to increase the length of intestine in juvenile rats. Methods: Fifty juvenile rats had double barrelled, blind loop ostomies created using an isolated segment of bowel. Our intestinal lengthening device was inserted into one of the loops and the second loop served as a control. Once the device was deployed, the experimental, control, and in situ segments of bowel were evaluated for length, weight, histology, and disaccharidase enzyme activity. Results: Mechanical tension increased intestinal length by 149%. The lengthened bowel also exhibited a greater total weight (218%), greater mucosal weight (122%), and increased protein mass (164%) compared with the control limb of bowel. Histologically, there was a markedly increased thickness of the muscularis propria in the lengthened bowel (200% increase compared with the control limb). Functionally, we found increased total disaccharidase activity in the lengthened bowel (between 47% and 350%, depending on the particular enzyme tested; p<0.01). Conclusion: Mechanical tension induces intestinal growth by increasing length, weight of the bowel and mucosa, and protein mass. Histological changes, such as increases in Paneth cells, suggest that increased proliferation and reorganisation of the mucosa and muscularis propria are a response to mechanical tension. Functionally, increased intestinal length corresponds with increased disaccharidase activity, thus implying potential increased absorptive capacity of the lengthened bowel.

Preoperative embolization as an adjunct to the operative management of mediastinal Castleman disease.

Castleman disease usually presents in children as a localized mass with prominent feeding vessels. The mainstay of treatment of Castleman disease is surgical resection; historically, resection is associated with excessive blood loss. These tumors are well known to have large feeding vessels and, thus, are amenable to preoperative arteriography with embolization. The authors present a case of Castleman disease treated with preoperative embolization as an adjunct to operative management.

Fine Mapping of Wilms’ Tumors With 16q Loss of Heterozygosity Localizes the Putative Tumor Suppressor Gene to a Region of 6.7 Megabases

Background:The aim of this study was to more precisely map the region of 16q loss of heterozygosity (LOH) in Wilms’ tumors and to examine the expression of putative tumor suppressor.Methods:We performed polymerase chain reaction–based LOH analysis on the 185 sample pairs from 21 to 80 megabases (Mb) on chromosome 16q. Expression of two candidate tumor suppressor genes located within the identified consensus region of 16q LOH was examined by immunohistochemistry.Results:We identified 16q LOH in 7 (4%) of 185 Wilms’ tumors not previously thought to demonstrate such genetic loss. The smallest common region of genetic loss was located between 67.3 and 74.0 Mb on chromosome 16. Within this 6.7-Mb region, there reside only three recognized tumor suppressor genes: E-cadherin, P-cadherin, and E2F4. E-cadherin demonstrates statistically significantly reduced expression in Wilms’ tumors with 16q LOH.Conclusions:We have localized the consensus region of 16q LOH in Wilms’ tumor to a 6.7-Mb locus and have identified three candidate Wilms’ tumor suppressor genes within this narrowed region. Our data support E-cadherin as a candidate tumor suppressor gene in Wilms’ tumor; however, further studies are needed to definitively prove its role as the tumor suppressor gene associated with 16q LOH.

Three phases of disaster relief in Haiti—pediatric surgical care on board the United States Naval Ship Comfort

BACKGROUND On January 12, 2010, Haiti experienced the western hemispheres worst-ever natural disaster. Within 24 hours, the United States Naval Ship Comfort received orders to respond, and a group of more than 500 physicians, nurses, and staff undertook the largest and most rapid triage and treatment since the inception of hospital ships. METHODS These data represent pediatric surgical patients treated aboard the United States Naval Ship Comfort between January 19 and February 27, 2010. Prospective databases managed by patient administration, radiology, blood bank, laboratory services, and surgical services were combined to create an overall patient care database that was retrospectively reviewed for this analysis. RESULTS Two hundred thirty-seven pediatric surgical patients were treated, representing 27% of the total patient population. These patients underwent a total of 213 operations composed of 243 unique procedures. Orthopedic procedures represented 71% of the total caseload. Patients returned to the operating room up to 11 times and required up to 28 days for completion of surgical management. CONCLUSIONS This represents the largest cohort of pediatric surgical patients in an earthquake response. Our analysis provides a model for anticipating surgical caseload, injury patterns, and duration of surgical course in preparing for future disaster response missions. Moreover, we propose a 3-phased response to disaster medicine that has not been previously described.

A Human Yeast Artificial Chromosome Containing the Multiple Endocrine Neoplasia Type 2B Ret Mutation Does Not Induce Medullary Thyroid Carcinoma but Does Support the Growth of Kidneys and Partially Rescues Enteric Nervous System Development in Ret-Deficient Mice

We generated a line of transgenic mice using a yeast artificial chromosome containing the Ret mutation responsible for the multiple endocrine neoplasia type 2B syndrome (MEN 2B). The resulting animals did not develop any of the expected neoplasms associated with MEN 2B. Transgenic animals were then bred with animals lacking murine Ret (Ret(M)) to further evaluate the function of human mutated Ret (Ret(H)(2B)) in the murine context. Whereas mice lacking Ret(M) exhibit intestinal aganglionosis and the absence of kidneys with other genitourinary anomalies, expression of the Ret(H)(2B) transgene in Ret(M)-deficient mice allowed significant renal development with a partial rescue of the enteric nervous system. These Ret(H)(2B)-positive/Ret(M)-deficient mice exhibit normal Ret expression and survive longer than Ret(M)-deficient mice, but still die at 3 to 5 days of age with evidence of enterocolitis. We conclude that the normal expression of a human Ret proto-oncogene with the MEN 2B mutation does not cause any features of MEN 2B in mice. Although the gene is normally expressed in the appropriate target tissues, there is incomplete phenotypic rescue in mice lacking murine Ret. These results suggest important interspecies differences between humans and mice in the function of the Ret oncogene.

Spinal abscess presenting as the initial symptom of Crohn’s disease

Dear Editor: Patients with Crohn’s disease commonly present with back pain from sacroiliitis and spondilitis. Spinal abscess from fistula formation is a rare complication from Crohn’s disease that can also present with back pain. We present a case of a patient presenting with a spinal abscess as his initial manifestation of Crohn’s disease. A 21-year-old Caucasian male with no previous medical history presented with 3 months of worsening back pain that he thought was the result of lifting a jet ski. He was initially diagnosed as having a bulging lumbar disc and was treated conservatively with analgesics. His pain, however, worsened and he was evaluated at a referring hospital where he was found to have abnormalities at L5 and the sacrum on imaging. An MRI showed an epidural abscess and he underwent a L3, L4, L5 laminectomy and coccygectomy to drain the presacral and epidural abscesses. Initial cultures demonstrated Streptococcus viridans and Bacteroides buccae. He underwent multiple debridements to the sacrum and lumbar spine, but progressively worsened with intractable pain, weakness in the lower extremities, decreased rectal tone, and urinary incontinence. A lower endoscopy was performed to investigate the colon as a possible source of the infection, but no communications or fistulae or obvious disease was identified. However, a presumptive diagnosis of Crohn’s disease was made based on random right colon biopsies. At this point, he was transferred to our institution for further evaluation and treatment. On admission the patient appeared emaciated and in poor health. His physical exam was remarkable for a normal abdominal exam, but decreased (4/5) strength in the quadriceps and biceps femoris and 1/5 strength of the flexors and extensors of the feet. He demonstrated normal sensation for deep pressure from L1 to S3, but had lost pinprick sensation from L5 to S3. There was a large open, granulating lumbar wound with foul drainage. His laboratory findings were significant for an elevated WBC of 13.7 with a left shift. MRI attained at this time demonstrated destruction and collapse of the L5 vertebral body with 85% retrolithesis of L4 and S1 and an inflammatory mass surrounding the sacrum with a diffuse phlegmon around the entire sacrum. He was placed on Unasyn and Cipro and after consultation with Orthopedics, Neurology, and Gastroenterology, was taken to the operating room for ileocecal and sigmoid resection, end colostomy, drainage of the retroperitoneal abscess, and anterior debridement of the L5 vertebral body. Two fistulae were evident upon entering the abdomen. The first, an ileosigmoid fistula, and the second a fistula form the involved are on the sigmoid down into the retroperitoneum. This second fistula coursed to the right side and then behind the mesorectum in the presacral space. The fistula then entered the lumbar vertebral bodies where orthopedic surgery performed the anterior debridement. An ileocecal resection with ileocolic anastomosis was performed along with a sigmoid resection with end colostomy. A wound V.A.C. was placed on the posterior wound to aid in drainage and closure. Pathology demonstrated active ileocecal Crohn’s disease. Intraoperative cultures grew Kiebsiella pneumoniae. Infectious disease consult optimized the antibiotics to include Zosyn and Cipro for 8 weeks. The patient recovered well from a GI standpoint and his WBC promptly returned to normal. He was placed Int J Colorectal Dis (2009) 24:601–602 DOI 10.1007/s00384-008-0599-5