Adipocyte Insulin-Mediated Glucose Transport: The Role of Myosin 1c, and a Method for \u3cem\u3ein vivo\u3c/em\u3e Investigation: A Dissertation

The importance of insulin delivery and action is best characterized in Type 2 Diabetes, a disease that is becoming a pandemic both nationally and globally. Obesity is a principal risk factor for Type 2 Diabetes, and adipocyte function abnormalities due to adipose hypertrophy and hyperplasia, have been linked to obesity. Numerous reports suggest that the intracellular and systemic consequences of adipocyte function abnormalities include adipocyte insulin resistance, enhanced production of free fatty acids, and production of inflammatory mediators. A hallmark of adipocyte insulin sensitivity is the stimulation of glucose transporter isoform 4 (GLUT4) trafficking events to promote glucose uptake. In the Type 2 diabetic and insulin resistant states the mechanism behind insulin-stimulated GLUT4 trafficking is compromised. Therefore, understanding the role of factors involved in glucose-uptake in adipose tissue is of great importance. Studies from our laboratory suggest an important role for the unconventional myosin, Myo1c, in promoting insulin-mediated glucose uptake in cultured adipocytes. Our observations suggest that depletion of Myo1c in cultured adipocytes results in a significant reduction in the ability of adipocytes to take up glucose following insulin treatment, suggesting Myo1c is required for insulin-mediated glucose uptake. A plausible mechanism by which Myo1c promotes glucose uptake in adipocytes has been suggested by further work from our laboratory in which expression of fluorescently-tagged Myo1c in cultured adipocytes induces significant membrane ruffling at the cell periphery, insulin-independent GLUT4 translocation to the cell periphery, and accumulation of GLUT4 in membrane ruffling regions. Taken together Myo1c seems to facilitate glucose uptake through remodeling of cortical actin. In the first part of this thesis I, in collaboration with others, uncovered a possible mechanism through which Myo1c regulates adipocyte membrane ruffling. Here we identified a novel protein complex in cultured adipocytes, comprising Myo1c and the mTOR binding partner, Rictor. Interestingly our studies in cultured adipocytes suggest that the Rictor-Myo1c complex is biochemically distinct from the Rictor-mTOR complex of mTORC2. Functionally, only depletion of Rictor but not Myo1c results in decreased Akt phosphorylation at serine 473, but depletion of either Rictor or Myo1c results in compromised cortical actin dynamic events. Furthermore we observed that whereas the overexpression of Myo1c in cultured adipocytes causes remarkable membrane ruffling, Rictor depletion in cells overexpressing Myo1c significantly reduces these ruffling events. Taken together our findings suggest that Myo1c, in conjunction with Rictor, modulates cortical actin remodeling events in cultured adipocytes. These findings have implications for GLUT4 trafficking as GLUT4 has been previously observed to accumulate in Myo1c-induced membrane ruffles prior to fusion with the plasma membrane. During our studies of adipocyte function we noticed that current siRNA electroporation methods present numerous limitations. To silence genes more effectively we employed a lentivirus-mediated shRNA delivery system, and to standardize this technology in cultured adipocytes we targeted Myo1c and MAP4K4. Using this technology we were able to achieve clear advantages over siRNA oligonucleotide electroporation techniques in stability and permanence of gene silencing. Furthermore we showed that the use of lentiviral vectors in cultured adipocytes did not affect insulin signaling or insulin-mediated glucose uptake events. Despite our inability to use lentiviral vectors to achieve gene silencing in mice we were able to achieve adipose tissue-specific gene silencing effects in mice following manipulation of the lentiviral conditional silencing vector, and then crossing resulting founders with aP2-Cre mice. Interestingly however, only founders from the MAP4K4 conditional shRNA vector, but not founders from the Myo1c conditional shRNA vector, showed gene knockdown, possibly due to position-effect variegation. Taken together, findings from these studies are important because they present an alternative means of achieving gene silencing in cultured adipocytes, with numerous advantages not offered by siRNA oligonucleotide electroporation methods. Furthermore, the in vivo, adipose tissue-specific RNAi studies offer a quick, inexpensive, and less technically challenging means of achieving adipose tissue-specific gene ablations relative to traditional gene knockout approaches

Trans-Tasman CGE Modelling: Some illustrative results from the J oani model

A two country multi-sectoral computable general equilibrium model for New 2.ealand and Australia is developed and applied to explore issues concerning the effects of the CER-induced tariff reductions and related topics. A comparison with a similar short-run exercise conducted in 1988 records minimal gains from a 1990 starting position as data shows little remaining protection to be removed. The removal of all protection arrangements with respect to imports from outside the CER region results in over 1 % extra GDP in the short run. However, this figure is doubled in both countries in the longer run as further gains from the re-allocation of capital resources amongst the sectors within each country are experienced. In interpreting the model results the critical nature of the assumptions incorporated within the model closure is discussed. Alternative assumptions addressing questions of inter-country capital mobility as well as exchange rate determination are examined

Tolerance to Non-Opioid Analgesics is Opioid Sensitive in the Nucleus Raphe Magnus

Repeated injection of opioid analgesics can lead to a progressive loss of effect. This phenomenon is known as tolerance. Several lines of investigations have shown that systemic, intraperitoneal administration or the microinjection of non-opioid analgesics, non-steroidal anti-inflammatory drugs (NSAIDs) into the midbrain periaqueductal gray matter induces antinociception with some effects of tolerance. Our recent study has revealed that microinjection of three drugs analgin, ketorolac, and xefocam into the central nucleus of amygdala produce tolerance to them and cross-tolerance to morphine. Here we report that repeated administrations of these NSAIDs into the nucleus raphe magnus (NRM) in the following 4 days result in progressively less antinociception compare to the saline control, i.e., tolerance develops to these drugs in male rats. Special control experiments showed that post-treatment with the μ-opioid antagonist naloxone into the NRM significantly decreased antinociceptive effects of NSAIDs on the first day of testing in the tail-flick (TF) reflex and hot plate (HP) latency tests. On the second day, naloxone generally had trend effects in both TF and HP tests and impeded the development of tolerance to the antinociceptive effect of non-opioid analgesics. These findings strongly support the suggestion of endogenous opioid involvement in NSAIDs antinociception and tolerance in the descending pain-control system. Moreover, repeated injections of NSAIDs progressively lead to tolerance to them, cross-tolerance to morphine, and the risk of a withdrawal syndrome. Therefore, these results are important for human medicine too

The Dynamics of the Developing Calcarenite Member of Pamutuan Formation In Cintaratu Area, Pangandaran, West Java

The investigated area is located in Cintaratu, Parigi District, Pangandaran Regency in the Southeastern part of Southern Mountain of West Java. The rock unit in the investigated area is described as the Calcarenite Member of the Pamutuan Formation. The present study applying detailed lithologic and paleontologic description reveals five lithologic units. Combining the field method with microscopic analysis leads to the verification of the age and environments of this carbonate rocks. The study concludes the development of reef complex in Early Mid Miocene, followed by the high energy of the dynamic environment producing the fragments of the lime clasts embedded in the packstone, indicates the deposition in the fore slope. The regression took place and moved the deposition environment to the low-energy condition at the back of the reef, producing bioclastic packstone containing larger foram. Rising seawater level amplified the dynamics of the seawater in mid-Middle Miocene. The formation of planktic packstone marked an initial transgression phase that was well-represented as regional transgressive in Southeast Asia. This phase continued until interbedded calcareous mudstone and sandstone were produced. Finally, in Late Mid Miocene the sea level regressive accumulated coarse materials of calcarenite grain size containing fragments of larger foram within shallower deposition environment. The NNW-SSE elongated distribution of five units of Calcerinite Member pointed out the configuration of Middle Miocene coastal line. In Early Middle Miocene, various carbonate rocks formation was primarily controlled by tectonic activities. Afterward, the rock deposition affected by the regional sea level rise and drop during Mid to Late Middle Miocene. The detailed description of the Calcarenite Member, therefore, contributed to the understanding of the paleogeography of Mid Miocene age and the dynamics of the environmental deposition

Peningkatan Penjualan Produk Rabbani melalui Penerapan Bisnis Intelijen dengan Metode Olap (Online Analytical Process)

Tahun 2015 reShare rabbani cipto-cirebon mengalami kondisi penjualan yang tidak stabil. Hal ini terjadi karena di daerah tersebut banyak berdiri toko-toko baru dari reShare lain. Akibatnyapenjualan produk rabbani mengalami penurunan omset pada 12 minggu terakhir ditahun 2015.Penelitian ini dilakukan menggunakan metode OLAP (Online Analytical Processing).Metode ini terdiri dari tiga tahapan operasi analisa dasar, diantaranya consolidation (roll-up), drill-down, slicing and dicing. Aplikasi bantu yang digunakan dalam penelitian ini yaituPowerOLAP versi 14. Penelitian ini diharapkan dapat membantu memberikan prediksi pada toko rabbani cipto-cirebon dalam meningkatkan penjualan produknya. Serta dapat membatu dalammemberikan inovasi baru pada produk rabbani agar banyak pengunjung datang. Sehingga dapat menghasilkan informasi yang tepat untuk membantu pihak manajemen dalam meningkatkanproduktifitas penjualan