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BACKGROUND There has been no previous study examining the phenomenon of suicide in older Asians in New Zealand. The aim of this study was to identify common factors and gain a better understanding of late-life suicide in Asian people living in New Zealand. METHODS New Zealand Coronial Services provided records of all closed cases of late-life suicides (age ≥ 65 years) between July 2007 and December 2012. Out of the total of 225 cases, 15 were recorded as being of Asian ethnicity. These cases were reviewed in their entirety using a qualitative thematic analysis approach. RESULTS Eight Asian men and seven Asian women completed suicide during this period. The majority (93%) lived with their families, and 80% were found by family after having completed suicide at home. Three main themes emerged from the thematic analysis: (i) suicide occurring in the context of a family; (ii) declining physical health; and (iii) a violent method of suicide. The role of the family has not been previously identified in other studies of late-life suicide in New Zealand. CONCLUSIONS Further research is needed to identify ways Asian families can access culturally appropriate and accessible support and mental health services for their older members at risk of suicide. There is also a need for developing post-suicide interventions specifically for Asian families. The findings of this study have added to the growing evidence of declining physical health acting as a drive for late-life suicide.

Analysis of embryonic fibroblasts from GFP reporter mice indicates that the fibroblast cell type harbors a large collection of developmentally and phenotypically heterogeneous subtypes. Some of these cells exhibit multipotency, whereas others do not. Multiparameter flow cytometry analysis shows that a large number of distinct populations of fibroblast-like cells can be found in cultures initiated from different embryonic organs, and cells sorted according to their surface phenotype typically retain their characteristics on continued propagation in culture. Similarly, surface phenotypes of individual cloned fibroblast-like cells exhibit significant variation. The fibroblast cell class appears to contain a very large number of denumerable subtypes.

There has been no previous study examining the phenomenon of suicide in older Asians in New Zealand. The aim of this study was to identify common factors and gain a better understanding of late-life suicide in Asian people living in New Zealand.New Zealand Coronial Services provided records of all closed cases of late-life suicides (age ≥ 65 years) between July 2007 and December 2012. Out of the total of 225 cases, 15 were recorded as being of Asian ethnicity. These cases were reviewed in their entirety using a qualitative thematic analysis approach.Eight Asian men and seven Asian women completed suicide during this period. The majority (93%) lived with their families, and 80% were found by family after having completed suicide at home. Three main themes emerged from the thematic analysis: (i) suicide occurring in the context of a family; (ii) declining physical health; and (iii) a violent method of suicide. The role of the family has not been previously identified in other studies of late-life suicide in New Zealand.Further research is needed to identify ways Asian families can access culturally appropriate and accessible support and mental health services for their older members at risk of suicide. There is also a need for developing post-suicide interventions specifically for Asian families. The findings of this study have added to the growing evidence of declining physical health acting as a drive for late-life suicide.

Efficient generation of competent vasculogenic cells is a critical challenge of human induced pluripotent stem (hiPS) cell-based regenerative medicine. Biologically relevant systems to assess functionality of the engineered vessels in vivo are equally important for such development. Here, we report a unique approach for the derivation of endothelial precursor cells from hiPS cells using a triple combination of selection markers—CD34, neuropilin 1, and human kinase insert domain-containing receptor—and an efficient 2D culture system for hiPS cell-derived endothelial precursor cell expansion. With these methods, we successfully generated endothelial cells (ECs) from hiPS cells obtained from healthy donors and formed stable functional blood vessels in vivo, lasting for 280 d in mice. In addition, we developed an approach to generate mesenchymal precursor cells (MPCs) from hiPS cells in parallel. Moreover, we successfully generated functional blood vessels in vivo using these ECs and MPCs derived from the same hiPS cell line. These data provide proof of the principle that autologous hiPS cell-derived vascular precursors can be used for in vivo applications, once safety and immunological issues of hiPS-based cellular therapy have been resolved. Additionally, the durability of hiPS-derived blood vessels in vivo demonstrates a potential translation of this approach in long-term vascularization for tissue engineering and treatment of vascular diseases. Of note, we have also successfully generated ECs and MPCs from type 1 diabetic patient-derived hiPS cell lines and use them to generate blood vessels in vivo, which is an important milestone toward clinical translation of this approach.

Rapid blood perfusion is critical for postimplantation survival of thick, prevascularized bioartificial tissues. Yet the mechanism by which implanted vascular networks inosculate, or anastomose, with the host vasculature has been unknown, making it difficult to develop optimized strategies for facilitating perfusion. Here we show that implanted vascular networks anastomose with host vessels through a previously unidentified process of “wrapping and tapping” between the engrafted endothelial cells (ECs) and the host vasculature. At the host-implant interface, implanted ECs first wrap around nearby host vessels and then cause basement membrane and pericyte reorganization and localized displacement of the underlying host endothelium. In this way, the implanted ECs replace segments of host vessels to divert blood flow to the developing implanted vascular network. The process is facilitated by high levels of matrix metalloproteinase-14 and matrix metalloproteinase-9 expressed by the wrapping ECs. These findings open the door to new strategies for improving perfusion of tissue grafts and may have implications for other physiologic and pathologic processes involving postnatal vasculogenesis.

Therapeutic revascularization with either exogenous angiogenic growth factors or vascular cells has yet to demonstrate efficacy in the clinic. Injection of angiogenic growth factors often produces unstable and abnormal blood vessels. Blood vascular networks derived from implanted endothelial cells persist only transiently due to the insufficient recruitment of perivascular cells. We hypothesize that a combination of the two approaches may act synergistically to yield a better result. To enhance the recruitment of perivascular cells, human umbilical vein endothelial cells were genetically modified to overexpress platelet-derived growth factor (PDGF)-BB. PDGF-BB overexpression promoted both proliferation and migration of perivascular precursor cells (10T1/2 cells) in vitro. When mock-infected endothelial cells were implanted alone in vivo, they formed transient blood vascular networks that regressed by day 30. PDGF-BB overexpression enhanced the survival of endothelial cells in vivo. However, the PDGF-BB-expressing vessel network failed to establish patent blood flow. Co-implantation of PDGF-BB-overexpressing endothelial cells with 10T1/2 cells paradoxically resulted in the rapid regression of the vascular networks in vivo. PDGF-BB stimulated the expression of both chemokine (C-C motif) ligand 2 (CCL2) and CCL7 in 10T1/2 cells and led to the increased accumulation of macrophages in vivo. These results suggest a potential negative interaction between angiogenic growth factors and vascular cells; their use in combination should be carefully tested in vivo for such opposing effects.

The rapidly changing field of gene therapy promises a number of innovative treatments for cancer patients. Advances in genetic modification of cancer and immune cells and the use of oncolytic viruses and bacteria have led to numerous clinical trials for cancer therapy, with several progressing to late-stage product development. At the time of this writing, no gene therapy product has been approved by the United States Food and Drug Administration (FDA). Some of the key scientific and regulatory issues include understanding of gene transfer vector biology, safety of vectors in vitro and in animal models, optimum gene transfer, long-term persistence or integration in the host, shedding of a virus and ability to maintain transgene expression in vivo for a desired period of time. Because of the biological complexity of these products, the FDA encourages a flexible, data-driven approach for preclinical safety testing programs. The clinical trial design should be based on the unique features of gene therapy products, and should ensure the safety of enrolled subjects. This article focuses on regulatory considerations for gene therapy product development and also discusses guidance documents that have been published by the FDA.

The field of regenerative medicine encompasses a breathtaking array of interdisciplinary scientific approaches with the promise of delivering future therapies to meet current unmet medical needs for patients. Increasingly more of these innovative products are being translated into human clinical trials in the United States, and general familiarity of the FDA is important to efficiently navigate the process. The basics of FDA history, organization, and processes are described herein for those new to clinical translation, with more detailed content added regarding approval pathways, regulations, guidances, and select special topics of relevance to regenerative medicine. In addition to the cumulative experience of previous products, the FDA regulatory approach to medical products evaluation includes an ongoing assessment of how the science of those products informs regulatory policy. FDA engages in ongoing dialogue with the scientific community and product sponsors to continue to develop science-based regulatory review policies that are robust and predictable in order to meet the needs of the challenging array of products that are on the horizon.

Purpose Circulating endothelial cells (CECs) and progenitor cells are currently evaluated as potential biomarkers of antiangiogenic therapy. CD146 is considered a panendothelial-specific marker, but its utility as a CEC marker in cancer patients remains unclear. Patients and Methods We analyzed the expression of CD146 on mononuclear blood cells, primary tissue endothelial cells, and malignant and normal tissues by flow cytometric and immunohistochemical analyses. Furthermore, we measured the circulation kinetics of CD146+ cells before, and then 3 and 12 days after vascular endothelial growth factor (VEGF) antibody blockade by bevacizumab infusion in rectal cancer patients enrolled in a phase I trial. Results In the peripheral blood of these cancer patients, over 90% of the CD146+ cells were CD45+ hematopoietic cells. CD146 expression was primarily detected on a subset of CD3+CD4+ lymphocytes, and was undetectable on CD34+CD133+CD45dim progenitor cells or CD31brightCD45− viable CECs. In contradistinction, CD146 was detectable in tissues on both cellular components of tumor vessel wall: CD31brightCD45− endothelial cells and α-SMA+ pericytes. Unlike viable CECs and progenitor cells, CD146+ cell concentration in the peripheral blood of cancer patients did not decrease during VEGF blockade. Conclusion CD146 is fairly homogeneously expressed on vascular endothelium but not on viable CECs or progenitor cells. The vast majority of CD146+ blood cells are lymphocytes, and VEGF blockade by bevacizumab did not significantly change their number in rectal cancer patients. These results underscore the need for further characterization and identification of new markers for CEC subpopulations for their development as biomarkers of antiangiogenic therapy.

The physical mechanism of the reflectance inversion phenomena observed in thin-film Pd on polyvinylidene fluoride (PVDF) optical hydrogen sensor structures (Pd/PVDF) upon exposure to hydrogen gas has been investigated. Based on experimental observations using sensors with or without a metallic back-surface coating, a theoretical model taking into account the infinite optical interreflections within the transparent polymer PVDF has been proposed. The understanding of the origin of the reflectance inversion is helpful in optimizing the design and performance of optical hydrogen sensors with Pd-layer active element structures.

The creation in vitro of vascularized skeletal muscle represents a first step to the engineering of more complex tissue architectures.

Scientists love a challenge. But for developers of cellular and gene therapy (CGT) products, understanding the scope of preclinical testing needed to support early-phase clinical trials can be daunting. In addition, these diverse and complex products often pose safety concerns that cannot be appropriately evaluated with traditional, standardized approaches to preclinical evaluation of small-molecule drugs and biologics. To help developers address these concerns, the U.S. Food and Drug Administration (FDA) has published a draft guidance for the preclinical assessment of investigational CGT products. This guidance clarifies current regulatory expectations for preclinical information used to support human clinical trials and license application. The draft guidance offers recommendations on the substance and scope of preclinical information needed to support clinical trials for multiple investigational product types, including cellular therapies, gene therapies, therapeutic vaccines, xenotransplantation products, and certain biologic-device combination products (such as cellularized scaffolds) regulated by the Office of Cellular, Tissue, and Gene Therapies (OCTGT). General areas of focus include selection of appropriate animal species and models of disease or injury and the conduct of proof-of-concept and toxicology studies. The guidance also highlights specific safety concerns for CGT products, such as tumorigenic potential for some cellular therapy products, and provides recommendations on approaches to their evaluation. Recommendations are presented in the context of designing preclinical programs that help define the risk-benefit profiles of investigational products in order to justify further testing in humans. This draft guidance, when finalized, will supersede the preclinical recommendations issued in a previous FDA guidance ([www.fda.gov/Drugs/ScienceResearch/ResearchAreas/ucm072987.htm][1]) and is not intended to provide a standardized or “cookbook” approach to the preclinical assessment of investigational CGT products. Rather, this guidance encourages a flexible and science-based approach to preclinical testing in which the kind, duration, and scope of testing are tailored for each product and target clinical indication. In recognizing the importance of product-specific, case-by-case application for the CGT products and the inherent challenges this poses, this guidance encourages early communication between product developers and the OCTGT pharmacology and toxicology review staff. In addition, the guidance emphasizes incorporation of the “3 Rs” principles (reduction, refinement, and replacement of animal use) in preclinical testing programs. Publication of this draft guidance signifies FDA’s continuing effort to engage CGT product developers to help facilitate the safe development of these emerging therapies. Draft Guidance for Industry: Preclinical Assessment of Investigational Cellular and Gene Therapy Products; [[Full Text] ][2] [1]: http://www.fda.gov/Drugs/ScienceResearch/ResearchAreas/ucm072987.htm [2]: http://www.fda.gov/downloads/BiologicsBloodVaccines/GuidanceComplianceRegulatoryInformation/Guidances/CellularandGeneTherapy/UCM329861.pdf

U.S. Food and Drug Administration applies regulatory flexibility to balance benefits and risks to subjects in cell-therapy clinical trials.

This chapter focuses on US Food and Drug Administration (FDA) regulatory considerations for stem/progenitor cell-based products (S/PCPs) intended to treat, mitigate or cure disease. Information critical to the regulatory decision-making process for determining the safety and efficacy of investigational S/PCPs is described. Included are sections that cover cellular product manufacturing and characterization, preclinical testing and clinical trial design. Increased regulatory complexity associated with stem/progenitor cell-based combination products is also discussed.

We have demonstrated a postprocessed complementary metal oxide semiconductor (CMOS) integrated circuit (IC) capable of on-chip magnetic separation, i.e., removing via magnetic forces the nonspecifically bound magnetic beads from the detection area on the surface of the chip. Initially, 4.5 mum wide superparamagnetic beads sedimenting out of solution due to gravity were attracted to the detection area by a magnetic concentration force generated by flowing current through a conductor embedded in the IC. After sedimentation, the magnetic beads that did not bind strongly to the functionalized surface of the IC through a specific biochemical complex were removed by a magnetic separation force generated by flowing current through another conductor placed laterally to the detection area. As the spherical bead pivoted on the surface of the chip, the lateral magnetic force was further amplified by mechanical leveraging, and 50 mA of current flowing through the separation conductor placed 18 mum away from the bead resulted in 7.5 pN of tensile force on the biomolecular tether immobilizing the bead. This force proved high enough to break nonspecific interactions while leaving specific antibody-antigen bonds intact. A sandwich capture immunoassay on purified human immunoglobulin G showed strong correlation with a control enzyme linked immunosorbent assay and a detection limit of 10 ngml or 70 pM. The beads bound to the detection area after on-chip magnetic separation were detected optically. To implement a fully integrated molecular diagnostics platform, the on-chip magnetic separation functionality presented in this work can be readily combine with state-of-the art CMOS-based magnetic bead detection technology.

A tiny footprint electrically probable single layer defocus monitor/test structure has been designed and tested to show sub-10nm resolution in electrical or electronic defocus monitoring. Electronic testing is a low-cost must have for on-chip production process monitoring which will become necessary for effective Design for Manufacturing. This programmable defocus monitor can be designed to pinch open at various levels of defocus by modifying four different layout parameters, CD, probe size, offset, and the number of rings. An array of these structures can be read as a series of opens and shorts, or 1s and 0s, to electronically extract defocus. One important feature of this defocus test structure is that it has an asymmetric response through focus, which translates to a high sensitivity to defocus at low defocus values or close to nominal conditions. Simulation and experimental results have shown good sensitivity for both on axis, tophat, and offaxis, quasar, illumination. This paper will present both simulation and experimental results that demonstrate the programmability and sensitivity of this test structure to defocus.

Vascular tissue engineering requires a ready source of endothelial cells and perivascular cells. Here, we evaluated human bone marrow–derived mesenchymal stem cells (hMSCs) for use as vascular progenitor cells in tissue engineering and regenerative medicine. hMSCs expressed a panel of smooth muscle markers in vitro including the cardiac/smooth muscle–specific transcription coactivator, myocardin. Cell-cell contact between endothelial cells and hMSCs up-regulated the transcription of myocardin. hMSCs efficiently stabilized nascent blood vessels in vivo by functioning as perivascular precursor cells. The engineered blood vessels derived from human umbilical cord vein endothelial cells and hMSCs remained stable and functional for more than 130 days in vivo. On the other hand, we could not detect differentiation of hMSCs to endothelial cells in vitro, and hMSCs by themselves could not form conduit for blood flow in vivo. Similar to normal perivascular cells, hMSC-derived perivascular cells contracted in response to endothelin-1 in vivo. In conclusion, hMSCs are perivascular cell precursors and may serve as an attractive source of cells for use in vascular tissue engineering and for the study of perivascular cell differentiation.

Tissue engineering has attracted wide interest as a potential method to alleviate the shortage of transplantable organs (1). To date, almost all of the successfully engineered tissues/organs have relatively thin and/or avascular structures [e.g., skin (2), cartilage (3), and bladder (4)], where postimplantation vascularization from the host (angiogenesis) is sufficient to meet the implant's demand for oxygen and nutrients. Vascularization remains a critical obstacle impeding attempts to engineer thicker, metabolically demanding organs, such as heart and liver. One approach in vascularizing an engineered tissue is to add the cellular components of blood vessels (endothelial and perivascular cells) directly to the tissue-engineered construct. We have shown that coimplanting endothelial cells and perivascular cells in a scaffold in vivo can lead to the formation of a vascular network that anastomoses to the host circulatory system. The engineered vessels are stable and functional, and they persist for more than 1 year in vivo. This approach may potentially lead to the creation of a well-vascularized-engineered tissue.

Tissue engineering has attracted wide interest as a potential method to alleviate the shortage of transplantable organs (1). To date, almost all of the successfully engineered tissues/organs have relatively thin and/or avascular structures [e.g., skin (2), cartilage (3), and bladder (4)], where postimplantation vascularization from the host (angiogenesis) is sufficient to meet the implant's demand for oxygen and nutrients. Vascularization remains a critical obstacle impeding attempts to engineer thicker, metabolically demanding organs, such as heart and liver. One approach in vascularizing an engineered tissue is to add the cellular components of blood vessels (endothelial and perivascular cells) directly to the tissue-engineered construct. We have shown that coimplanting endothelial cells and perivascular cells in a scaffold in vivo can lead to the formation of a vascular network that anastomoses to the host circulatory system. The engineered vessels are stable and functional, and they persist for more than 1 year in vivo. This approach may potentially lead to the creation of a well-vascularized-engineered tissue.

Tissue engineering requires formation of a de novo stable vascular network. Because of their ability to proliferate, differentiate into endothelial cells, and form new vessels, blood-derived endothelial progenitor cells (EPCs) are attractive source of cells for use in engineering blood vessels. However, the durability and function of EPC-derived vessels implanted in vivo are unclear. To this end, we directly compared formation and functions of tissue-engineered blood vessels generated by peripheral blood– and umbilical cord blood–derived EPCs in a model of in vivo vasculogenesis. We found that adult peripheral blood EPCs form blood vessels that are unstable and regress within 3 weeks. In contrast, umbilical cord blood EPCs form normal-functioning blood vessels that last for more than 4 months. These vessels exhibit normal blood flow, perm-selectivity to macromolecules, and induction of leukocyte-endothelial interactions in response to cytokine activation similar to normal vessels. Thus, umbilical cord blood EPCs hold great therapeutic potential, and their use should be pursued for vascular engineering.

A database and data analysis strategy is proposed for multi-designer test chips that involve a wide array of different test structures aimed at process characterization. The database described in this paper has been custom built for the multi-student FLCC testchip that has six contributing students and over 15,000 individually probably transistors/test structures. It has an interface with the Parametric Yield Simulator (PYS) that drives simulation parameters and automatically populates the database with simulation results of each test structure at the specified process conditions. A well-designed database forces structure into measurement and design related data, but includes enough flexibility as to adapt to different types of test structures and experiments. The database is split into four separate sections that store description of test structures, simulated results, experiment results, and process conditions. All data is centrally located and web accessible for easy access from any computer with Internet access. Simulation results can be uploaded from the server running the PYS, experimental results can be uploaded directly from the lab and data can be compared and queried by all users of the database. Data analysis strategies can be compared and reused as queries and data analysis results can be shared among users through the website. Queries can be saved, loaded, rated, and reused, so even novice SQL users can utilize advanced queries. Advanced queries form the basis of a strategy that first identifies good process monitors based on simulation results and then uses them to extract process conditions from electrical measurements via an iterative process. This paper describes strategies that can be used to help facilitate collaboration and hence leverage the benefits of combining multiple sets of test structures from different designers on one chip.

Hb Lepore-Hollandia, identified in a Thai patient, was found to be due to a new cross-over between IVS1-nt 42 and nt 56 of the delta- and beta-globin genes. This differs from a previous case which crossed-over between codon 22 and IVS1-nt 16. Two independent homologous unequal cross-over events account for these two Lepore-Hollandia genes.

The construction of stable blood vessels is a fundamental challenge for tissue engineering in regenerative medicine. Although certain genes can be introduced into vascular cells to enhance their survival and proliferation, these manipulations may be oncogenic. We show here that a network of long-lasting blood vessels can be formed in mice by co-implantation of vascular endothelial cells and mesenchymal precursor cells, by-passing the need for risky genetic manipulations. These networks are stable and functional for one year in vivo.

The construction of stable blood vessels is a fundamental challenge for tissue engineering in regenerative medicine. Although certain genes can be introduced into vascular cells to enhance their survival and proliferation, these manipulations may be oncogenic. We show here that a network of long-lasting blood vessels can be formed in mice by co-implantation of vascular endothelial cells and mesenchymal precursor cells, by-passing the need for risky genetic manipulations. These networks are stable and functional for one year in vivo.

Human embryonic stem cell (hESC) lines provide not only an unique system for studying human embryonic development, but also have enormous potential as a source of therapeutic tissues, including those of vascular and hematopoietic lineages. However, it is difficult to generate large-scale of relatively pure population of specific lineage cells derived from hESCs. Here, we describe a 2-dimensional (2-D) culture system for hESC differentiation to generate hematopoietic and endothelial cells in vitro without embryoid body (EB) formation (3-D). To initial differentiation, hESCs (H1 and H9 cell lines) were placed in serum-containing media for 9–10 days without supplement of additional growth factors. Gene expression analysis of 2-D differentiation indicated that Oct-4, a pluripotent gene, was decreased, while genes for GATA-2 and CD31, were increased, suggesting that differentiated hESCs contain hematopoietic and endothelial progenitors. Comparing with 3-D EB formation, 2-D differentiation was able to generate robust differentiated cells (5 to 10-fold higher). To generate a reasonable pure population of hematopoietic and endothelial precursors, CD34+ cells prior to hematopoietic commitment were isolated from 2-D differentiation by immuno-magnetic beads and cultured in the presence of hematopoietic growth factors and endothelial growth factors. After 7–10 days culture of CD34+ cells, both suspension and adherent cells emerged. The number of suspension and adherent cells was dependent on the presence of hematopoietic growth factors or/and endothelial growth factors. The suspension cells expressed hematopoietic marker, CD45, and gave rise to multi-lineage hematopoietic colonies on methylcellulose cultures. The adherent cells expressed endothelial markers, such as CD31 and Ve-cadherin, and took up Dill-labeled ac-LDL. The adherent cells rapidly formed capillary-like structures on Matrigel cultures, while the suspension cells would not. While the adherent cells were implanted into mice, they formed functional blood vessels in vivo. Our culture system paves the road for further large-scale manufacture of hematopoietic and endothelial cells derived from human ES cells. Our studies demonstrated also that CD34+ cells derived from human ES cells contain hematopoietic and endothelial progenitors.