Endocytic recycling protein EHD1 regulates primary cilia morphogenesis and SHH signaling during neural tube development.

Members of the four-member C-terminal EPS15-Homology Domain-containing (EHD) protein family play crucial roles in endocytic recycling of cell surface receptors from endosomes to the plasma membrane. In this study, we show that Ehd1 gene knockout in mice on a predominantly B6 background is embryonic lethal. Ehd1-null embryos die at mid-gestation with a failure to complete key developmental processes including neural tube closure, axial turning and patterning of the neural tube. We found that Ehd1-null embryos display short and stubby cilia on the developing neuroepithelium at embryonic day 9.5 (E9.5). Loss of EHD1 also deregulates the ciliary SHH signaling with Ehd1-null embryos displaying features indicative of increased SHH signaling, including a significant downregulation in the formation of the GLI3 repressor and increase in the ventral neuronal markers specified by SHH. Using Ehd1-null MEFS we found that EHD1 protein co-localizes with the SHH receptor Smoothened in the primary cilia upon ligand stimulation. Under the same conditions, EHD1 was shown to co-traffic with Smoothened into the developing primary cilia and we identify EHD1 as a direct binding partner of Smoothened. Overall, our studies identify the endocytic recycling regulator EHD1 as a novel regulator of the primary cilium-associated trafficking of Smoothened and Hedgehog signaling.

Novel treatment for mantle cell lymphoma including therapy-resistant tumor by NF-κB and mTOR dual-targeting approach.

Mantle cell lymphoma (MCL) is one of the most aggressive B-cell non-Hodgkin lymphomas with a median survival of approximately five years. Currently, there is no curative therapy available for refractory MCL because of relapse from therapy-resistant tumor cells. The NF-κB and mTOR pathways are constitutively active in refractory MCL leading to increased proliferation and survival. Targeting these pathways is an ideal strategy to improve therapy for refractory MCL. Therefore, we investigated the in vitro and in vivo antilymphoma activity and associated molecular mechanism of action of a novel compound, 13-197, a quinoxaline analog that specifically perturbs IκB kinase (IKK) ß, a key regulator of the NF-κB pathway. 13-197 decreased the proliferation and induced apoptosis in MCL cells including therapy-resistant cells compared with control cells. Furthermore, we observed downregulation of IκBα phosphorylation and inhibition of NF-κB nuclear translocation by 13-197 in MCL cells. In addition, NF-κB-regulated genes such as cyclin D1, Bcl-XL, and Mcl-1 were downregulated in 13-197-treated cells. In addition, 13-197 inhibited the phosphorylation of S6K and 4E-BP1, the downstream molecules of mTOR pathway that are also activated in refractory MCL. Further, 13-197 reduced the tumor burden in vivo in the kidney, liver, and lungs of therapy-resistant MCL-bearing nonobese diabetic severe-combined immunodeficient (NOD/SCID) mice compared with vehicle-treated mice; indeed, 13-197 significantly increased the survival of MCL-transplanted mice. Together, results suggest that 13-197 as a single agent disrupts the NF-κB and mTOR pathways leading to suppression of proliferation and increased apoptosis in malignant MCL cells including reduction in tumor burden in mice.

Introduction of ultrasound into gross anatomy curriculum: perceptions of medical students.

BACKGROUND: The exposure to ultrasound technology during medical school education is highly variable across institutions. OBJECTIVES: The objectives of this study were to assess medical students' perceptions of ultrasound use to teach Gross Anatomy along with traditional teaching methods, and determine their ability to identify sonographic anatomy after focused didactic sessions. METHODS: Prospective observational study. Phase I of the study included three focused ultrasound didactic sessions integrated into Gross Anatomy curriculum. During Phase II, first-year medical students completed a questionnaire. RESULTS: One hundred nine subjects participated in this study; 96% (95% confidence interval [CI] 92-99%) agreed that ultrasound-based teaching increased students' knowledge of anatomy acquired through traditional teaching methods. Ninety-two percent (95% CI 87-97%) indicated that ultrasound-based teaching increases confidence to perform invasive procedures in the future. Ninety-one percent (95% CI 85-96%) believed that it is feasible to integrate ultrasound into the current Anatomy curriculum. Ninety-eight percent (95% CI 95-100%) of medical students accurately identified vascular structures on ultrasound images of normal anatomy of the neck. On a scale of 1 to 10, the average confidence level reported in interpreting the images was 7.4 (95% CI 7.1-7.7). Overall, 94% (95% CI 91-99%) accurately answered questions about ultrasound fundamentals and sonographic anatomy. CONCLUSIONS: The majority of medical students believed that it is feasible and beneficial to use ultrasound in conjunction with traditional teaching methods to teach Gross Anatomy. Medical students were very accurate in identifying sonographic vascular anatomy of the neck after brief didactic sessions.

The influence of surgical demonstrations during an anatomy course on the perceptions of first-year medical students toward surgeons and a surgical career.

OBJECTIVE: We previously have demonstrated the educational benefits of surgical demonstrations to first-year medical students. The aim of this current study was to analyze the influence of these demonstrations on the perceptions of students toward surgeons and a possible career in surgery. METHODS: A faculty member from the Department of Surgery provided an instruction on pancreatic malignancies and management to first-year medical students during their gross anatomy course. After this instruction, using a lightly embalmed cadaver, the clinically relevant anatomy was detailed and a pancreaticoduodenectomy was performed on the cadaver. Immediately after the demonstration, a brief survey was conducted to obtain feedback from the students about the experience. RESULTS: A total of 170 students over 2 years returned the survey for a response rate of 69%. The demonstration provided 77% of students with a favorable impression of surgeons, and 90% of the students felt that this exposure gave them an understanding of the knowledge, skills, and qualities needed to become a surgeon. Additionally, 57% of respondents stated that watching the demonstration increased the likelihood of them pursuing a surgical career. For the 67% of students who were considering a surgery career, the demonstration reinforced their interest; however, for the students who were not interested in surgery, the demonstration did not alter their opinion. CONCLUSION: The results of this study showed that surgical demonstrations to first-year medical students can influence their perceptions favorably about surgeons and a surgical career. This interaction provided students with information and motivation to pursue a career in surgery and also may counteract any negative stereotypes of the field that first-year students may have had.

The endocytic recycling regulator EHD1 is essential for spermatogenesis and male fertility in mice.

BACKGROUND: The C-terminal Eps15 homology domain-containing protein 1 (EHD1) is ubiquitously expressed and regulates the endocytic trafficking and recycling of membrane components and several transmembrane receptors. To elucidate the function of EHD1 in mammalian development, we generated Ehd1-/- mice using a Cre/loxP system. RESULTS: Both male and female Ehd1-/- mice survived at sub-Mendelian ratios. A proportion of Ehd1-/- mice were viable and showed smaller size at birth, which continued into adulthood. Ehd1-/- adult males were infertile and displayed decreased testis size, whereas Ehd1-/- females were fertile. In situ hybridization and immunohistochemistry of developing wildtype mouse testes revealed EHD1 expression in most cells of the seminiferous epithelia. Histopathology revealed abnormal spermatogenesis in the seminiferous tubules and the absence of mature spermatozoa in the epididymides of Ehd1-/- males. Seminiferous tubules showed disruption of the normal spermatogenic cycle with abnormal acrosomal development on round spermatids, clumping of acrosomes, misaligned spermatids and the absence of normal elongated spermatids in Ehd1-/- males. Light and electron microscopy analyses indicated that elongated spermatids were abnormally phagocytosed by Sertoli cells in Ehd1-/- mice. CONCLUSIONS: Contrary to a previous report, these results demonstrate an important role for EHD1 in pre- and post-natal development with a specific role in spermatogenesis.

An experience in surgical anatomy to provide first-year medical students with an early exposure to general surgery: a pilot study.

INTRODUCTION: The level of interest expressed by medical students toward the field of general surgery has decreased. The aims of this study were to (1) describe an educational scheme in surgical anatomy that increases interaction between practicing surgeons and first-year medical students and (2) garner feedback and opinions of these medical students from a pilot study of this educational experience. MATERIALS AND METHODS: A faculty member from the Department of Surgery provided a review of pancreatic malignancies and its management to first-year medical students during their anatomy course. Then, using a cadaver, the clinically relevant anatomy was detailed, and a pancreaticoduodenectomy was performed with the help of student volunteers. A 7-question survey using the 5-point Likert response scale ranging from "strongly agree" to "strongly disagree" was used to obtain feedback from the students. RESULTS: A total of 145 responses (of 205) were collected for a response rate of 70.38%. Most students (99%) felt that this type of surgical demonstration during the anatomy course was extremely beneficial. The students also felt that this approach improved their understanding of the relevant anatomy and its clinical importance. The survey also demonstrated that most students would like these surgical demonstrations to be repeated in future. Less than 1% of the students did not find these demonstrations beneficial. CONCLUSIONS: The results of this study demonstrate the benefit of surgical demonstrations by surgical faculty to first-year medical students. These findings have led to the incorporation of this educational scheme into the medical school anatomy curriculum on a regular basis at our University.

Developing an online prematriculation orientation program and its relation to student performance in the first class taken in medical school.

BACKGROUND: In 2004 the University of Nebraska College of Medicine developed an online prematriculation program, Fast Start, to introduce students to the environment and expectations in medical school. PURPOSE: This quantitative study was conducted to determine whether using Fast Start correlated with performance in the gross anatomy course. METHODS: A hierarchical regression analysis was used to correlate grades in gross anatomy with a set of common prediction variables and a variable for use of Fast Start. RESULTS: The results showed that the predictive power of the full model, including the Fast Start variable, was slightly stronger than for the reduced model. A separate model verified the absence of an interaction between Fast Start use and prior academic ability. CONCLUSIONS: The online Fast Start program provided an efficient and effective method of delivering a prematriculation student orientation experience; its use was associated with marginally improved performance in a medical school course.

Comparison of phenotypic and functional dendritic cells derived from human umbilical cord blood and peripheral blood mononuclear cells.

Dendritic cells (DC) are important accessory cells that are capable of initiating an immune response. Generation of functional DC has potential clinical use in treating diseases such as cancer. In this report, we have demonstrated the generation of functional DC from mononuclear cells isolated from human umbilical cord blood cells (UCBC) and peripheral blood cells (PBC) using a defined medium Prime Complete Growth Medium (PCGM) (GenePrime LLC, Gaithersburg, MD). DC generated using PCGM showed the typical phenotype of DC as determined by flow cytometry and electron microscopy. Further analysis of the DC using confocal microscopy showed localization of the antigen and major histocompatibility complex (MHC) molecules in the cytoplasm 3-5 days following tumor antigen loading into DC. Subsequently, the tumor antigen-MHC complex was localized on the surface of DC. DC generated from UCBC or PBC also increased (p < 0.001) the allogeneic mixed lymphocyte reaction, confirming their immune accessory functions compared to a control mixed lymphocyte reaction (MLR) without DC added. Interestingly, DC generated using PCGM medium also significantly enhanced the hematopoietic colony (CFU-C)-forming ability. Furthermore, addition of 5% DC derived from cord blood loaded with tumor antigen also significantly (p < 0.001) increased peripheral and cord blood-derived antigen-specific cytotoxic T lymphocyte (CTL)-mediated killing of human leukemic cells (K562) and breast cancer cells (MDA-231). Thus, these results show that functional DC generated from cord blood using a defined medium are a useful source of accessory cells for augmenting CTL-mediated cytotoxicity and have potential use in cellular therapy for human leukemia and breast cancer.