MD/PhD Program Copyright (c) 2013 University of Massachusetts Medical School All rights reserved. http://escholarship.umassmed.edu/gsbs_mdphd Recent documents in MD/PhD Program en-us Wed, 13 Feb 2013 16:32:58 PST 3600 Leptin treatment confers clinical benefit at multiple stages of virally induced type 1 diabetes in BB rats http://escholarship.umassmed.edu/gsbs_mdphd/19 http://escholarship.umassmed.edu/gsbs_mdphd/19 Mon, 20 Sep 2010 06:17:43 PDT The adipokine, leptin, regulates blood glucose and the insulin secretory function of beta cells, while also modulating immune cell function. We hypothesized that the dual effects of leptin may prevent or suppress the autoreactive destruction of beta cells in a virally induced rodent model of type 1 diabetes. Nearly 100% of weanling BBDR rats treated with the combination of an innate immune system activator, polyinosinic:polycytidylic acid (pIC), and Kilham rat virus (KRV) become diabetic within a predictable time frame. We utilized this model to test the efficacy of leptin in preventing diabetes onset, remitting new onset disease, and preventing autoimmune recurrence in diabetic rats transplanted with syngeneic islet grafts. High doses of leptin delivered via an adenovirus vector (AdLeptin) or alzet pump prevented diabetes in>90% of rats treated with pIC+KRV. The serum hyperleptinemia generated by this treatment was associated with decreased body weight, decreased non-fasting serum insulin levels, and lack of islet insulitis in leptin-treated rats. In new onset diabetics, hyperleptinemia prevented rapid weight loss and diabetic ketoacidosis, and temporarily restored euglycemia. Leptin treatment also prolonged the survival of syngeneic islets transplanted into diabetic BBDR rats. In diverse therapeutic settings, we found leptin treatment to have significant beneficial effects in modulating virally induced diabetes. These findings merit further evaluation of leptin as a potential adjunct therapeutic agent for treatment of human type 1 diabetes.

]]> Annie J. Kruger et al. Diabetes Mellitus, Type 1 Leptin Rats, Inbred BB Islets of Langerhans Transplantation Enterohemorrhagic E. coli requires N-WASP for efficient type III translocation but not for EspFU-mediated actin pedestal formation http://escholarship.umassmed.edu/gsbs_mdphd/18 http://escholarship.umassmed.edu/gsbs_mdphd/18 Thu, 16 Sep 2010 08:55:00 PDT Upon infection of mammalian cells, enterohemorrhagic E. coli (EHEC) O157:H7 utilizes a type III secretion system to translocate the effectors Tir and EspF(U) (aka TccP) that trigger the formation of F-actin-rich 'pedestals' beneath bound bacteria. EspF(U) is localized to the plasma membrane by Tir and binds the nucleation-promoting factor N-WASP, which in turn activates the Arp2/3 actin assembly complex. Although N-WASP has been shown to be required for EHEC pedestal formation, the precise steps in the process that it influences have not been determined. We found that N-WASP and actin assembly promote EHEC-mediated translocation of Tir and EspF(U) into mammalian host cells. When we utilized the related pathogen enteropathogenic E. coli to enhance type III translocation of EHEC Tir and EspF(U), we found surprisingly that actin pedestals were generated on N-WASP-deficient cells. Similar to pedestal formation on wild type cells, Tir and EspF(U) were the only bacterial effectors required for pedestal formation, and the EspF(U) sequences required to interact with N-WASP were found to also be essential to stimulate this alternate actin assembly pathway. In the absence of N-WASP, the Arp2/3 complex was both recruited to sites of bacterial attachment and required for actin assembly. Our results indicate that actin assembly facilitates type III translocation, and reveal that EspF(U), presumably by recruiting an alternate host factor that can signal to the Arp2/3 complex, exhibits remarkable versatility in its strategies for stimulating actin polymerization.

]]> Didier F. Vingadassalom et al. Enterohemorrhagic Escherichia coli Wiskott-Aldrich Syndrome Protein, Neuronal Carrier Proteins Escherichia coli Proteins Protein Transport Actins T-bet knockout prevents Helicobacter felis-induced gastric cancer http://escholarship.umassmed.edu/gsbs_mdphd/17 http://escholarship.umassmed.edu/gsbs_mdphd/17 Fri, 10 Sep 2010 08:36:03 PDT Helicobacter infection is the primary risk factor for gastric cancer, with the cytokine environment within the gastric mucosa the strongest predictor of disease risk. Elevated TNF-alpha, IL-1beta, and low IL-10 are associated with the highest risk. In this study, we used C57BL/6 mice to identify T-bet as a central regulator of the cytokine environment during Helicobacter felis infection. We infected male and female C57BL/6 and C57BL/6-T-bet knockout (KO) litter mates with H. felis and examined the bacterial colonization, immune response, and mucosal damage at varying time points. T-bet KO mice maintained infection for 15 mo at similar levels to wild-type mice. Infection and immune response did not differ between male and female mice. Despite sustained infection, T-bet KO mice respond with a blunted Th1 response associated with preservation of parietal and chief cells and protection from the development of gastric cancer. Unexpectedly, T-bet KO mice develop a gastric environment that would not be expected based on the phenotype of T-bet KO CD4 cells alone. T-bet KO mice respond to H. felis infection with a markedly blunted IL-1beta and TNF-alpha and elevated IL-10 levels. Activity of this one master regulator modulates the expression of the key gastric mucosal cytokines associated with gastric cancer and may be a target for therapy to restore immune balance clinically in patients at risk for gastric cancer.

]]> Calin Stoicov et al. Adenocarcinoma Animals Female Gastric Mucosa Genetic Predisposition to Disease Helicobacter Infections Helicobacter felis Interleukin-1beta Male Mice Mice, Inbred C57BL Mice, Knockout Stomach Neoplasms T-Box Domain Proteins Tumor Necrosis Factor-alpha How the study of Helicobacter infection can contribute to the understanding of carcinoma development http://escholarship.umassmed.edu/gsbs_mdphd/16 http://escholarship.umassmed.edu/gsbs_mdphd/16 Fri, 10 Sep 2010 08:36:02 PDT The inflammatory environment dramatically impacts the formation of cancer at many levels, acting on the stem cell to foster the initiation of cancer all the way through its contribution to metastatic disease. Using Helicobacter-induced gastric cancer as an example, it can be seen that, early on, chronic inflammation exhausts tissue stem cells, forcing the remaining stem cells to work overtime and calling in replacement cells from marrow sources. Marrow-derived stromal cells orchestrate growth and remodelling through secreted factors and cell-cell communication. Once cancer is present, the inflammatory environment is responsible for the continued growth signals to the cancer stem cells and to the stromal cells which become a vital part of the cancer niche as well as the pre-metastatic niche which will effectively lure cancer cells into peripheral organs for distant growth. This understanding of the inflammatory environment and its many effects on cancer throughout its natural history provides intervention targets directed at the unique aspects of cancer behaviour.

]]> Calin Stoicov et al. Carcinoma Helicobacter Helicobacter Infections Humans Inflammation The intrinsically disordered cytoplasmic domain of the T cell receptor zeta chain binds to the nef protein of simian immunodeficiency virus without a disorder-to-order transition. http://escholarship.umassmed.edu/gsbs_mdphd/15 http://escholarship.umassmed.edu/gsbs_mdphd/15 Mon, 10 May 2010 12:46:38 PDT Intrinsically disordered proteins are thought to undergo coupled binding and folding upon interaction with their folded partners. In this study, we investigate whether binding of the intrinsically disordered T cell receptor zeta cytoplasmic tail to the well-folded simian immunodeficiency virus Nef core domain is accompanied by a disorder-to-order transition. We show that zeta forms a 1:1 complex with Nef and remains unfolded in the complex. Thus, our findings oppose the generally accepted view of the behavior of intrinsically disordered proteins and provide new evidence of the existence of specific interactions for unfolded protein molecules.

]]> Alexander B. Sigalov et al. Binding Sites Cytoplasm Dimerization Electrophoresis, Polyacrylamide Gel Gene Products, nef Protein Folding Protein Structure, Tertiary Receptors, Antigen, T-Cell Simian immunodeficiency virus Viral pathogenesis, modulation of immune receptor signaling and treatment. http://escholarship.umassmed.edu/gsbs_mdphd/14 http://escholarship.umassmed.edu/gsbs_mdphd/14 Mon, 10 May 2010 12:46:37 PDT During the co-evolution of viruses and their hosts, the latter have equipped themselves with an elaborate immune system to defend themselves from the invading viruses. In order to establish a successful infection, replicate and persist in the host, viruses have evolved numerous strategies to counter and evade host antiviral immune responses as well as exploit them for productive viral replication. These strategies include those that target immune receptor transmembrane signaling. Uncovering the exact molecular mechanisms underlying these critical points in viral pathogenesis will not only help us understand strategies used by viruses to escape from the host immune surveillance but also reveal new therapeutic targets for antiviral as well as immunomodulatory therapy. In this chapter, based on our current understanding of transmembrane signal transduction mediated by multichain immune recognition receptors (MIRRs) and the results of sequence analysis, we discuss the MIRR-targetingviral strategies of immune evasion and suggest their possible mechanisms that, in turn, reveal new points of antiviral intervention. We also show how two unrelated enveloped viruses, human immunodeficiency virus and human cytomegalovirus, use a similar mechanism to modulate the host immune response mediated by two functionally different MIRRs-T-cell antigen receptor and natural killer cell receptor, NKp30. This suggests that it is very likely that similar general mechanisms can be or are used by other viral and possibly nonviral pathogens.

]]> Walter Minsub Kim et al. Amino Acid Sequence Animals Humans Molecular Sequence Data Receptors, Immunologic Signal Transduction Virus Diseases Virus Internalization Virus Replication Viruses Pseudo-merohedral twinning and noncrystallographic symmetry in orthorhombic crystals of SIVmac239 Nef core domain bound to different-length TCRzeta fragments. http://escholarship.umassmed.edu/gsbs_mdphd/13 http://escholarship.umassmed.edu/gsbs_mdphd/13 Mon, 10 May 2010 12:46:36 PDT HIV/SIV Nef mediates many cellular processes through interactions with various cytoplasmic and membrane-associated host proteins, including the signalling zeta subunit of the T-cell receptor (TCRzeta). Here, the crystallization strategy, methods and refinement procedures used to solve the structures of the core domain of the SIVmac239 isolate of Nef (Nef(core)) in complex with two different TCRzeta fragments are described. The structure of SIVmac239 Nef(core) bound to the longer TCRzeta polypeptide (Leu51-Asp93) was determined to 3.7 A resolution (R(work) = 28.7%) in the tetragonal space group P4(3)2(1)2. The structure of SIVmac239 Nef(core) in complex with the shorter TCRzeta polypeptide (Ala63-Arg80) was determined to 2.05 A resolution (R(work) = 17.0%), but only after the detection of nearly perfect pseudo-merohedral crystal twinning and proper assignment of the orthorhombic space group P2(1)2(1)2(1). The reduction in crystal space-group symmetry induced by the truncated TCRzeta polypeptide appears to be caused by the rearrangement of crystal-contact hydrogen-bonding networks and the substitution of crystallographic symmetry operations by similar noncrystallographic symmetry (NCS) operations. The combination of NCS rotations that were nearly parallel to the twin operation (k, h, -l) and a and b unit-cell parameters that were nearly identical predisposed the P2(1)2(1)2(1) crystal form to pseudo-merohedral twinning.

]]> Walter Minsub Kim et al. Amino Acid Sequence Crystallography, X-Ray Gene Products, nef Hydrogen Bonding Ligands Models, Molecular Molecular Sequence Data Peptide Fragments Protein Binding Protein Interaction Domains and Motifs Protein Structure, Quaternary Receptors, Antigen, T-Cell Simian immunodeficiency virus The role of innate immune pathways in type 1 diabetes pathogenesis. http://escholarship.umassmed.edu/gsbs_mdphd/12 http://escholarship.umassmed.edu/gsbs_mdphd/12 Mon, 10 May 2010 12:46:35 PDT PURPOSE OF REVIEW: Type 1 diabetes (T1D) is an autoimmune disease typically believed to result from malfunctions in adaptive immune response signaling which result in activation of self-reactive T cells. However, recent research has indicated components of the innate immune response as having a key role in the initiation of the autoimmune process of T1D. This review will highlight recent studies which examined the role of innate immune response signaling and the connections to T1D pathogenesis.

RECENT FINDINGS: Investigations indicate that components of innate immunity, including inflammation and Toll-like receptor signaling, are involved in pancreatic islet infiltration and insulitis. Recent studies examining the role of viral infections in T1D development also implicate innate immune response signaling in disease pathogenesis.

SUMMARY: Current research indicates that components of innate immune response signaling are involved in the initiation of the autoimmune process which results in the eventual destruction of beta cells during T1D pathogenesis. Continuing efforts by researchers to uncover the molecular pathways of innate immunity linked to T1D development could potentially lead to therapeutics capable of preventing and curing the autoimmune disease.

]]> Steven C. Pino et al. Diabetes Mellitus, Type 1 Immunity, Innate Depletion of the programmed death-1 receptor completely reverses established clonal anergy in CD4(+) T lymphocytes via an interleukin-2-dependent mechanism http://escholarship.umassmed.edu/gsbs_mdphd/11 http://escholarship.umassmed.edu/gsbs_mdphd/11 Mon, 14 Sep 2009 12:12:57 PDT Recent studies have implicated the cell surface receptor Programmed Death-1 (PD-1) in numerous models of T cell anergy, though the specific mechanisms by which the PD-1 signal maintains tolerance is not clear. We demonstrate that the depletion of PD-1 with siRNA results in a complete reversal of clonal anergy in the A.E7 T cell model, suggesting that the mechanism by which PD-1 maintains the anergic phenotype is a T-cell-intrinsic phenomenon, and not one dependent on other cell populations in vivo. We have also shown that the neutralization of IL-2 during restimulation abrogates the effect of PD-1 depletion, suggesting that tolerance mediated by PD-1 is wholly IL-2 dependent, and likewise intrinsic to the tolerized cells.

]]> Kenneth D. Bishop et al. Animals Antigens Antigens, Surface Apoptosis Regulatory Proteins inhibitors Base Sequence CD4-Positive T-Lymphocytes Cell Line Cell Proliferation Clonal Anergy Columbidae Cytochromes c DNA Primers Interleukin-2 Mice Mice, Inbred BALB C Phenotype RNA, Messenger RNA, Small Interfering Signal Transduction Transfection Transplantation Tolerance Dynein light intermediate chain 1 is required for progress through the spindle assembly checkpoint http://escholarship.umassmed.edu/gsbs_mdphd/10 http://escholarship.umassmed.edu/gsbs_mdphd/10 Mon, 14 Sep 2009 12:12:55 PDT The spindle assembly checkpoint monitors microtubule attachment to kinetochores and tension across sister kinetochores to ensure accurate division of chromosomes between daughter cells. Cytoplasmic dynein functions in the checkpoint, apparently by moving critical checkpoint components off kinetochores. The dynein subunit required for this function is unknown. Here we show that human cells depleted of dynein light intermediate chain 1 (LIC1) delay in metaphase with increased interkinetochore distances; dynein remains intact, localised and functional. The checkpoint proteins Mad1/2 and Zw10 localise to kinetochores under full tension, whereas BubR1 is diminished at kinetochores. Metaphase delay and increased interkinetochore distances are suppressed by depletion of Mad1, Mad2 or BubR1 or by re-expression of wtLIC1 or a Cdk1 site phosphomimetic LIC1 mutant, but not Cdk1-phosphorylation-deficient LIC1. When the checkpoint is activated by microtubule depolymerisation, Mad1/2 and BubR1 localise to kinetochores. We conclude that a Cdk1 phosphorylated form of LIC1 is required to remove Mad1/2 and Zw10 but not BubR1 from kinetochores during spindle assembly checkpoint silencing.

]]> Mylavarapu V. S. Sivaram et al. Calcium-Binding Proteins Cell Cycle Proteins Cells, Cultured Dynein ATPase Hela Cells Humans Kinetochores Metaphase Mitotic Spindle Apparatus Phosphorylation Protein-Serine-Threonine Kinases Repressor Proteins BRCA1 does not paint the inactive X to localize XIST RNA but may contribute to broad changes in cancer that impact XIST and Xi heterochromatin http://escholarship.umassmed.edu/gsbs_mdphd/9 http://escholarship.umassmed.edu/gsbs_mdphd/9 Mon, 14 Sep 2009 12:12:54 PDT The BRCA1 tumor suppressor involved in breast and ovarian cancer is linked to several fundamental cell regulatory processes. Recently, it was reported that BRCA1 supports localization of XIST RNA to the inactive X chromosome (Xi) in women. The apparent cytological overlap between BRCA1 and XIST RNA across the Xi raised the possibility a direct role of BRCA1 in localizing XIST. We report here that BRCA1 does not paint the Xi or XIST territory, as do markers of Xi facultative heterochromatin. A smaller BRCA1 accumulation abuts Xi, although this is not exclusive to Xi. In BRCA1 depleted normal and tumor cells, or BRCA1 reconstituted cells, BRCA1 status does not closely correlate with XIST localization, however in a BRCA1 inducible system over-expression correlated strongly with enhanced XIST expression. We confirm frequent loss of an Xi in tumor cells. In addition to mitotic loss of Xi, we find XIST RNA expression or localization frequently become compromised in cultured breast cancer cells, suggesting Xi heterochromatin may not be fully maintained. We demonstrate that complex epigenetic differences between tumor cell subpopulations can have striking effects on XIST transcription, accumulation, and localization, but this does not strictly correlate with BRCA1. Although BRCA1 can have indirect effects that impact XIST, our results do not indicate a direct and specific role in XIST RNA regulation. Rather, regulatory factors such as BRCA1 that have broad effects on chromatin or gene regulation can impact XIST RNA and the Xi. We provide preliminary evidence that this may occur as part of a wider failure of heterochromatin maintenance in some cancers.

]]> Gayle Jeannette Pageau et al. BRCA1 Protein Breast Neoplasms Cell Line Cell Line, Tumor Chromosomes, Human, X Female Heterochromatin Humans In Situ Hybridization, Fluorescence RNA Interference RNA, Untranslated X Chromosome Inactivation Neuroprotective effect of hyperbaric oxygen therapy monitored by MR-imaging after embolic stroke in rats http://escholarship.umassmed.edu/gsbs_mdphd/8 http://escholarship.umassmed.edu/gsbs_mdphd/8 Mon, 14 Sep 2009 12:12:53 PDT The potential neuroprotective effects of hyperbaric oxygen (HBO) were tested in an embolic model of focal cerebral ischemia with partially spontaneous reperfusion. Rats (n = 10) were subjected to embolic middle cerebral artery occlusion (MCAO) and diffusion weighted MRI (DWI) was performed at baseline, 1, 3, and 6 h after MCAO to determine the ADC viability threshold yielding the lesion volumes that best approximated the 2,3,5-triphenyltetrazolium chloride (TTC) infarct volumes at 24 h (experiment 1). For assessment of neuroprotective effects, rats were treated with 100% oxygen at 2.5 atmospheres absolute (ATA, n = 15) or normobaric room air (n = 15) for 60 min beginning 180 min after MCAO (experiment 2). DWI-, perfusion (PWI)- and T2-weighted MRI (T2WI) started within 0.5 h after MCAO and was continued 5 h, 24 h (PWI and T2WI only), and 168 h (T2WI only). Infarct volume was calculated based on TTC-staining at 24 h (experiment 1) or 168 h (experiment 2) post-MCAO. ADC-lesion evolution was maximal between 3 and 6 h. In experiment 2, the relative regional cerebral blood volume (rCBV) of both groups showed similar incomplete spontaneous reperfusion in the ischemic core. HBO reduced infarct volume to 145.3 +/- 39.6 mm3 vs. 202.5 +/- 58.3 mm3 (control, P = 0.029). As shown by MRI and TTC, HBO treatment demonstrated significant neuroprotection at 5 h after embolic focal cerebral ischemia that lasted for 168 h.

]]> Nils Henninger et al. Animals Arterial Occlusive Diseases Brain Infarction Brain Ischemia Cerebral Arterial Diseases Cerebral Cortex Embolism *Hyperbaric Oxygenation Magnetic Resonance Imaging Male Middle Cerebral Artery Neuroprotective Agents Oxygen Rats Rats, Wistar Stroke Survival Analysis Time Factors A new model of thromboembolic stroke in the posterior circulation of the rat http://escholarship.umassmed.edu/gsbs_mdphd/7 http://escholarship.umassmed.edu/gsbs_mdphd/7 Mon, 14 Sep 2009 12:12:51 PDT The prognosis of vertebrobasilar occlusion is grave and therapeutic options are limited. The aim of the present study was to develop a new model of embolic hindbrain ischemia in the rat that closely resembles the clinical situation and that can be used to study pathophysiology and treatment options. After thoracotomy in 20 male Wistar rats, 15 animals received an injection of in vitro prepared autologous blood clots into the left vertebral artery. Five animals without clot injection served as controls. Neurological deficits were assessed in all animals 2 h after embolism. After 2 h, five animals were sacrificed to measure cerebral blood flow (CBF) by iodo-antipyridine autoradiography, and to calculate early cerebellar swelling by comparison of both hemispheres in brain slices. In these animals, autoradiography revealed ipsilesional brain swelling and significantly reduced blood flow values relative to the contralateral (unaffected) structures. Immunohistology showed the typical pattern of focal cerebral ischemia in the brain stem and/or cerebellum in 7 of 10 animals allowed to recover to 24 h. Hence, successful thromboembolism was achieved in 12 of 15 animals (80%). With this novel model, the pathophysiology and potential treatments of posterior circulation stroke can be investigated.

]]> Nils Henninger et al. Animals Autoradiography Brain Edema Cerebral Infarction Cerebrovascular Circulation Disease Models, Animal Immunohistochemistry Intracranial Embolism Male Rats Rats, Wistar Stroke Thromboembolism Impaired spatial learning in a novel rat model of mild cerebral concussion injury http://escholarship.umassmed.edu/gsbs_mdphd/6 http://escholarship.umassmed.edu/gsbs_mdphd/6 Mon, 14 Sep 2009 12:12:50 PDT The aim of the present study was to develop a model of mild traumatic brain injury in the rat that mimics human concussive brain injury suitable to study pathophysiology and potential treatments. 34 male Wistar rats received a closed head trauma (TBI) and 30 animals served as controls (CON). Immediately following trauma, animals lost their muscle tone and righting reflex response, recovering from the latter within 11.4 +/- 8.2 min. Corneal reflex and whisker responses returned within 4.5 +/- 3.0 min and 6.1 +/- 2.9 min, respectively. The impact resulted in a short transient decrease of pO2 (P < 0.001), increase in mean arterial blood pressure (P = 0.026), and a reduction of heart rate (P < 0.01). Serial MRI did not show any abnormalities across the entire cerebrum on diffusion, T1, T2, and T2*-weighted images at all investigated time points. TBI animals needed significantly longer to locate the hidden platform in a Morris water maze and spent less time in the training quadrant than controls. TBI led to a significant neuronal loss in frontal cortex (P < 0.001), as well as hippocampal CA3 (P = 0.017) and CA1 (P = 0.002) at 9 days after the trauma; however, cytoskeletal architecture was preserved as indicated by normal betaAPP- and MAP-2 staining. We present a unique, noninvasive rat model of mild closed head trauma with characteristics of human concussion injury, including brief loss of consciousness, cognitive impairment, and minor brain injury.

]]> Nils Henninger et al. Analysis of Variance Animals Behavior, Animal Blood Pressure Brain Brain Concussion Cell Count Cell Death Disease Models, Animal Heart Rate Immunohistochemistry Learning Disorders Magnetic Resonance Imaging Male Maze Learning Microtubule-Associated Proteins Rats Rats, Wistar Spatial Behavior Time Factors Effects of hypoxia, hyperoxia, and hypercapnia on baseline and stimulus-evoked BOLD, CBF, and CMRO2 in spontaneously breathing animals http://escholarship.umassmed.edu/gsbs_mdphd/5 http://escholarship.umassmed.edu/gsbs_mdphd/5 Mon, 14 Sep 2009 12:12:48 PDT Functional magnetic resonance imaging (fMRI) was used to investigate the effects of inspired hypoxic, hyperoxic, and hypercapnic gases on baseline and stimulus-evoked changes in blood oxygenation level-dependent (BOLD) signals, cerebral blood flow (CBF), and the cerebral metabolic rate of oxygen (CMRO2) in spontaneously breathing rats under isoflurane anesthesia. Each animal was subjected to a baseline period of six inspired gas conditions (9% O2, 12% O2, 21% O2, 100% O2, 5% CO2, and 10% CO2) followed by a superimposed period of forepaw stimulation. Significant stimulus-evoked fMRI responses were found in the primary somatosensory cortices. Relative fMRI responses to forepaw stimulation varied across gas conditions and were dependent on baseline physiology, whereas absolute fMRI responses were similar across moderate gas conditions (12% O2, 21% O2 100% O2, and 5% CO2) and were relatively independent of baseline physiology. Consistent with data obtained using well-established techniques, baseline and stimulus-evoked CMRO2 were invariant across moderate physiological perturbations thereby supporting a CMRO2-fMRI technique for non-invasive CMRO2 measurement. However, under 9% O2 and 10% CO2, stimulus-evoked CBF and BOLD were substantially reduced and the CMRO2 formalism appeared invalid, likely due to attenuated neurovascular coupling and/or a failure of the model under extreme physiological perturbations. These findings demonstrate that absolute fMRI measurements help distinguish neural from non-neural contributions to the fMRI signals and may lend a more accurate measure of brain activity during states of altered basal physiology. Moreover, since numerous pharmacologic agents, pathophysiological states, and psychiatric conditions alter baseline physiology independent of neural activity, these results have implications for neuroimaging studies using relative fMRI changes to map brain activity.

]]> Kenneth M. Sicard et al. Animals Blood Flow Velocity Blood Volume Brain Carbon Dioxide Electric Stimulation Energy Metabolism Forelimb Heart Rate Homeostasis *Image Enhancement *Image Processing, Computer-Assisted *Magnetic Resonance Imaging Male Oxygen Oxygen Consumption Rats Rats, Sprague-Dawley Somatosensory Cortex Vascular Resistance Imaging oxygen consumption in forepaw somatosensory stimulation in rats under isoflurane anesthesia http://escholarship.umassmed.edu/gsbs_mdphd/4 http://escholarship.umassmed.edu/gsbs_mdphd/4 Mon, 14 Sep 2009 12:12:47 PDT The cerebral metabolic rate of oxygen (CMRO2) was dynamically evaluated on a pixel-by-pixel basis in isoflurane-anesthetized and spontaneously breathing rats following graded electrical somatosensory forepaw stimulations (4, 6, and 8 mA). In contrast to alpha-chloralose, which is the most widely used anesthetic in forepaw-stimulation fMRI studies of rats under mechanical ventilation, isoflurane (1.1-1.2%) provided a stable anesthesia level over a prolonged period, without the need to adjust the ventilation volume/rate or sample blood gases. Combined cerebral blood flow signals (CBF) and blood oxygenation level-dependent (BOLD) fMRI signals were simultaneously measured with the use of a multislice continuous arterial spin labeling (CASL) technique (two-coil setup). CMRO2 was calculated using the biophysical BOLD model of Ogawa et al. (Proc Natl Acad Sci USA 1992;89:5951-5955). The stimulus-evoked BOLD percent changes at 4, 6, and 8 A were, respectively, 0.5% +/- 0.2%, 1.4% +/- 0.3%, and 2.0% +/- 0.3% (mean +/- SD, N = 6). The CBF percent changes were 23% +/- 6%, 58% +/- 9%, and 87% +/- 14%. The CMRO2 percent changes were 14% +/- 4%, 24% +/- 6%, and 43% +/- 11%. BOLD, CBF, and CMRO2 activations were localized to the forepaw somatosensory cortices without evidence of plateau for oxygen consumption, indicative of partial coupling of CBF and CMRO2. This study describes a useful forepaw-stimulation model for fMRI, and demonstrate that CMRO2 changes can be dynamically imaged on a pixel-by-pixel basis in a single setting with high spatiotemporal resolution.

]]> Zhaohui M. Liu et al. Anesthetics, Inhalation Animals Brain Brain Mapping Cerebrovascular Circulation Electric Stimulation Forelimb Isoflurane Magnetic Resonance Imaging Male Oxygen Rats Rats, Sprague-Dawley Respiration Respiration, Artificial Spin Labels Regional cerebral blood flow and BOLD responses in conscious and anesthetized rats under basal and hypercapnic conditions: implications for functional MRI studies http://escholarship.umassmed.edu/gsbs_mdphd/3 http://escholarship.umassmed.edu/gsbs_mdphd/3 Mon, 14 Sep 2009 12:12:46 PDT Anesthetics, widely used in magnetic resonance imaging (MRI) studies to avoid movement artifacts, could have profound effects on cerebral blood flow (CBF) and cerebrovascular coupling relative to the awake condition. Quantitative CBF and tissue oxygenation (blood oxygen level-dependent [BOLD]) were measured, using the continuous arterial-spin-labeling technique with echo-planar-imaging acquisition, in awake and anesthetized (2% isoflurane) rats under basal and hypercapnic conditions. All basal blood gases were within physiologic ranges. Blood pressure, respiration, and heart rates were within physiologic ranges in the awake condition but were depressed under anesthesia (P < 0.05). Regional CBF was heterogeneous with whole-brain CBF values of 0.86 +/- 0.25 and 1.27 +/- 0.29 mL. g-1. min-1 under awake and anesthetized conditions, respectively. Surprisingly, CBF was markedly higher (20% to 70% across different brain conditions) under isoflurane-anesthetized condition compared with the awake state (P < 0.01). Hypercapnia decreased pH, and increased Pco(2) and Po(2). During 5% CO(2) challenge, under awake and anesthetized conditions, respectively, CBF increased 51 +/- 11% and 25 +/- 4%, and BOLD increased 7.3 +/- 0.7% and 5.4 +/- 0.4%. During 10% CO(2) challenge, CBF increased 158 +/- 28% and 47 +/- 11%, and BOLD increased 12.5 +/- 0.9% and 7.2 +/- 0.5%. Since CBF and BOLD responses were substantially higher under awake condition whereas blood gases were not statistically different, it was concluded that cerebrovascular reactivity was suppressed by anesthetics. This study also shows that perfusion and perfusion-based functional MRI can be performed in awake animals.

]]> Kenneth M. Sicard et al. Anesthesia Animals Carbon Dioxide Cerebrovascular Circulation Consciousness Hypercapnia *Magnetic Resonance Imaging Oxygen Rats Rats, Sprague-Dawley Spin Labels DNA vaccines: vector design, delivery, and antigen presentation http://escholarship.umassmed.edu/gsbs_mdphd/2 http://escholarship.umassmed.edu/gsbs_mdphd/2 Mon, 14 Sep 2009 12:12:45 PDT Inoculations with antigen-expressing plasmid DNAs (DNA vaccines) in the production of protective immune responses. Since the initial development of DNA vaccines more than 5 years ago, major strides have been made in the design of efficient vaccine vectors and in the process of vaccine delivery. However, many questions remain regarding the mechanism of cellular transfection and in the development of immune responses. This review addresses functional aspects of DNA vaccines, including vector design and delivery, as well as cellular transfection and antigen presentation.

]]> David Marc Feltquate Animals *Antigen Presentation Drug Delivery Systems Drug Design Genetic Engineering Genetic Vectors Humans Vaccines, DNA Wilson disease in 1998: genetic, diagnostic and therapeutic aspects http://escholarship.umassmed.edu/gsbs_mdphd/1 http://escholarship.umassmed.edu/gsbs_mdphd/1 Mon, 14 Sep 2009 12:12:43 PDT John L. Gollan et al. *Hepatolenticular Degeneration Humans