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Papers by Michael Michalkiewicz

Warfarin is one of the most frequently prescribed drugs used to prevent blood clotting. However, ... more Warfarin is one of the most frequently prescribed drugs used to prevent blood clotting. However, warfarin's therapeutic window is narrow (2.0 Objective: The Aurora Health Care (AHC) Research Institute and Zilber School of Public Health, University of Wisconsin-Milwaukee are conducting a multi-phase retrospective EMR-based longitudinal anticoagulation clinical database project to study anticoagulation dosing algorithms. Methods: Steps in the database development include: requirements analysis (identification of clinical variables); Data collection, quality control and integration process; and modeling and simulation analysis. Data sources included two EMR systems. A one-way honest broker was used to protect patient confidentiality. Results: The initial database contains over 110,000 unique anticoagulation patients from 15 inpatient facilities across south-eastern Wisconsin from 2002-2009. Age, race and geographical distributions of the cohort are consistent with the State of Wisc...

Transgenics in Endocrinology

In the early 1980s, Palmiter, Brinster, and their colleagues (1,2) reported dramatic effects of o... more In the early 1980s, Palmiter, Brinster, and their colleagues (1,2) reported dramatic effects of overexpression of a human or a rat growth hormone (GH) under control of a mouse metallothionein-I (MT) promoter in transgenic mice. Introduction of these gene constructs into the mouse genome produced striking acceleration of growth and a major increase in adult body size. Adult GH-transgenic mice are not obese, but may weigh twice as much as their normal siblings. Impressive alterations of the phenotype of these giant mice described by Palmiter et al. (1,2) and by other pioneers of this field (3) provided a demonstration of the enormous potential of transgenic technology, and undoubtedly contributed to the wide use of transgenic organisms in biological research, agriculture, and research targeted at developing gene therapies for various diseases.

International Journal of Poultry Science, 2008

Objective of the study is examine the effects of active immunization against chicken Vasoactive I... more Objective of the study is examine the effects of active immunization against chicken Vasoactive Intestinal Peptide (chVIP) on plasma prolactin (PRL) concentration, concentration of luteinizing hormone (LH) profile its interval and duration, Progesterone (P), Estradiol (E ß), intersequence pause days and egg 4 2 production in birds during the later stages of egg production from 48-72 weeks of egg lay in PB3 birds. Twenty-four PB3 birds of same age group were divided into 2 groups of 12 in each. Birds in control group were administered s/c with placebo. Equal volume of chVIP immunogenic protein was administered to treated group from 17 week of age to 36 weeks of age at an interval of 4 weeks. Hormonal profiles of th immunized and control birds were quantified at weekly intervals from 48th to 72nd weeks of age in both the groups for prolactin, LH, estradiol, progesterone. Egg production and pause days were recorded in both the groups. At 59th weeks of age, blood samples from chVIP immunized and control birds were obtained every 4 h for 48 h to study the surges of LH. In immunized PB3 birds (against ch VIP) plasma PRL concentration was lower (p<0.01) with high concentrations of E ß, P , LH and its 4 h LH surges in plasma. Significantly 2 4 (p<0.01) higher egg production (13.62%) and less pause days were observed in chVIP immunized birds. It is mainly attributed due to low PRL concentration, associated with high concentrations of LH (with regular interval and duration of LH surges), E ß and P concentration required for egg formation and subsequent 2 4 ovulation. In conclusion, chVIP immunization advanced the LH surges, for release of matured oocyte, egg formation and egg lay enabled the birds to lay eggs at regular intervals due to active immunization against chVIP. Results indicate that control of chVIP may lead to more egg production with shorter duration of LH surges.

Proceedings of the National Academy of Sciences, 2000

Exogenous neuropeptide Y (NPY) reduces experimental anxiety in a wide range of animal models. The... more Exogenous neuropeptide Y (NPY) reduces experimental anxiety in a wide range of animal models. The generation of an NPY-transgenic rat has provided a unique model to examine the role of endogenous NPY in control of stress and anxiety-related behaviors using paradigms previously used by pharmacological studies. Locomotor activity and baseline behavior on the elevated plus maze were normal in transgenic subjects. Two robust phenotypic traits were observed. ( i ) Transgenic subjects showed a markedly attenuated sensitivity to behavioral consequences of stress, in that they were insensitive to the normal anxiogenic-like effect of restraint stress on the elevated plus maze and displayed absent fear suppression of behavior in a punished drinking test. ( ii ) A selective impairment of spatial memory acquisition was found in the Morris water maze. Control experiments suggest these traits to be independent. These phenotypic traits were accompanied by an overexpression of prepro-NPY mRNA and N...

Permissions: Requests for permissions to reproduce figures, tables, or portions of articles origi... more Permissions: Requests for permissions to reproduce figures, tables, or portions of articles originally published in Hypertension can be obtained via RightsLink, a service of the Copyright Clearance Center, not the Editorial Office. Once the online version of the published article for which permission is being requested is located, click Request Permissions in the middle column of the Web page under Services. Further information about this process is available in the Permissions and Rights Question and Answer document. Reprints: Information about reprints can be found online at:

Neuropeptide Y is a potent inhibitory neurotransmitter expressed in the central neurons that cont... more Neuropeptide Y is a potent inhibitory neurotransmitter expressed in the central neurons that control blood pressure. NO also serves as an inhibitory neurotransmitter, and its deficit causes sympathetic overactivity, which then contributes to hypertension. This study tested the hypothesis that neuropeptide Y functions as a central neurotransmitter to lower blood pressure, therefore its increased signaling ameliorates hypertension induced by NO deficiency. Conscious neuropeptide Y transgenic male rats, overexpressing the peptide under its natural promoter, and nontransgenic littermates (controls) were used in this study. Neuropeptide Y, Y1 receptor antagonist BIBP3226, or vehicle (saline) were administered continuously for 14 days into the cerebral lateral ventricle in unrestrained animals using osmotic pumps. Blood pressure was measured by radiotelemetry. Compared with control animals, transgenic overexpression of neuropeptide Y significantly ameliorated (by 9.7 1.5 mm Hg) NO deficie...

American Journal of Physiology-Endocrinology and Metabolism, 1993

It has been suggested that thyroid blood flow is regulated by both sympathetic and parasympatheti... more It has been suggested that thyroid blood flow is regulated by both sympathetic and parasympathetic nerves. The purpose of our experiments was to study the role of neuropeptide Y (NPY) in the sympathetic neural control of thyroid blood flow. Sympathetic nerve fibers to the thyroid contain both norepinephrine (NE) and NPY. Therefore, NE (15 nmol iv bolus) and NPY (12 or 1.7 nmol/kg body wt iv infusion; 4 min) were administered to anesthetized male rats (250–300 g) either alone or together, with or without an alpha-adrenergic receptor blocker (phentolamine; 10 mg/kg body wt iv bolus). Experiments were also performed in which the cervical sympathetic trunks were stimulated (30 Hz, 10 V; 0.5 ms; 2 min) with or without phentolamine. Thyroid blood flow was monitored continuously by laser-Doppler blood flowmetry. Results are expressed as thyroid vascular conductance (TVC). NE or NPY at both doses decreased TVC relative to that in control saline-infused rats (P < 0.05). No potentiation of...

NPY, previously known as a sympathetic vasoconstricting co-transmitter, recently has also emerged... more NPY, previously known as a sympathetic vasoconstricting co-transmitter, recently has also emerged as an important vascular growth factor, stimulating proliferation of vascular smooth muscle and endothelial cells (ECs). Both in vitro and in vivo, NPY stimulates angiogenesis starting at sub-picomolar, non-vasoconstrictive concentrations, and with maximal effects similar to those of other known angiogenic factors e.g. vascular endothelial growth factor (VEGF). To determine if NPY is a physiological mediator in vivo, we studied 1 ) ischemic angiogenesis in a rodent model of femoral artery occlusion 2) vascular development in rats over-expressing NPY gene; 3) changes in angiogenesis with aging. Hindlimb ischemia increased local NPY release and shifted NPY receptor (R) expression from predominantly Yl to the Y2R type, and up-regulated dipeptidyl peptidase IV (DPPIV, peptidase forming Y2/YS agonist). Increasing local NPY levels 2-fold with a slow-release pellet (Il.g/14 days, below arterial occlusion), additionally induced YSR mRNA, and stimulated capillary angiogenesis and collateral vessel growth - leading to restoration of blood flow and contractile function of ischemic muscles. These effects were markedly reduced in Y2R-/- mice, as was NPY-induced aortic sprouting ex vivo; the latter was also reduced by anti-VEGF antibody and completely abolished in endothelial nitric oxide synthase (eNOS)-/- mice. The role of NPY in vascular development was further revealed by the severe impairment of spontaneous aortic sprouting in NPY-/- mice and changes in vascular density of non-ischemic muscles: reduction in Y2-/- mice and marked increase in NPY-Tg rats. With aging, in mice, NPY-induced angiogenesis decreased together with Y2 and DPPIV expression, and in human ECs, the proliferative effects of NPY declined too, similarly to those of VEGF or basic fibroblast growth factor (bFGF). Since NPY-induced EC proliferation was similarly blocked by anti-VEGF and anti-FGF antibodies in young and old patients, this suggests that age-induced impaired activity of the NPY-Y2R system could lead to lower secretion of these growth factors, in addition to decreased responsiveness of old vessels to VEGF and bFGF. Taken together, our studies indicate that NPY is a physiological neurogenic factor playing an important role ip revascularization of ischemic tissues and in age-related changes in vascular development. NPY's angiogenic activity requires Y2Rs and eNOS, and involves release of VEGF and bFGF. Thus, sympathetic nerve activity, via NPY, may be an important upstream mechanism triggering a cascade of molecular events leading to vascular remodeling during tissue ischemia and growth.

Neuroscience, 2002

öFunctional studies in epileptic tissue indicate that neuropeptide Y and some of its peptide anal... more öFunctional studies in epileptic tissue indicate that neuropeptide Y and some of its peptide analogs potently inhibit seizure activity. We investigated seizure susceptibility in transgenic rats overexpressing the rat neuropeptide Y gene under the control of its natural promoter. Seizures were induced in adult transgenic male rats and their wild-type littermates by i.c.v. injection of 0.3 Wg kainic acid or by electrical kindling of the dorsal hippocampus. Transgenic rats showed a signi¢cant reduction in the number and duration of electroencephalographic seizures induced by kainate by 30% and 55% respectively (P 6 0.05 and 0.01). Transgenic rats were also less susceptible to epileptogenesis than wild-type littermates as demonstrated by a 65% increase in the number of electrical stimuli required to induce stage 5 seizures (P 6 0.01). This phenotype was associated with a strong and speci¢c expression of neuropeptide Y mRNA in area CA1, a brain area involved in the seizure network. We conclude that endogenous neuropeptide Y overexpression in the rat hippocampus is associated with inhibition of seizures and epileptogenesis suggesting that this system may be a valuable target for developing novel antiepileptic treatments.

Physiological Genomics, 2004

A single point mutation in a novel immune-associated nucleotide gene 5 ( Ian5) coincides with sev... more A single point mutation in a novel immune-associated nucleotide gene 5 ( Ian5) coincides with severe T cell lymphopenia in BB rats. We used a transgenic rescue approach in lymphopenic BB-derived congenic F344. lyp/ lyp rats to determine whether this mutation is responsible for lymphopenia and to establish the functional importance of this novel gene. A 150-kb P1 artificial chromosome (PAC) transgene harboring a wild-type allele of the rat Ian5 gene restored Ian5 transcript and protein levels, completely rescuing the T cell lymphopenia in the F344. lyp/ lyp rats. This successful complementation provides direct functional evidence that the Ian5 gene product is essential for maintaining normal T cell levels. It also demonstrates that transgenic rescue in the rat is a practical and definitive method for revealing the function of a novel gene.

Molecular and Cellular Neuroscience, 1992

Both neuropeptide Y (NPY) and vasoactive intestinal peptide (VIP) are present in thyroid nerves a... more Both neuropeptide Y (NPY) and vasoactive intestinal peptide (VIP) are present in thyroid nerves and have been shown to alter thyroid activity. The present study was conducted to determine whether hypo- or hyperthyroidism is associated with changes in the expression of the mRNAs for these neuropeptides in the major ganglia which supply nerves to the thyroid or within the thyroid gland itself. Hypo- or hyperthyroid conditions were induced by the administration of propylthiouracil (PTU) or thyroxine (T(4)), respectively, for 6 days. Control rats received vehicle injections. Total RNA from superior cervical ganglia (SCG), local thyroid ganglia, thyroid gland, and selected other tissues was extracted and mRNA levels were analyzed using Northern blot procedures. No significant changes in preproNPY or precursor VIP mRNA levels were detected in the SCG or the local thyroid ganglia in response to PTU or T(4) treatment. However, PTU treatment was associated with an increase in preproNPY mRNA levels in the thyroid gland itself. These results indicate that changes within the thyroid axis in response to these hypo- and hyperthyroid conditions do not include alterations in steady-state preproNPY or precursor VIP mRNA concentrations in the major ganglia which supply nerves to the thyroid gland. However, intrathyroidal preproNPY mRNA levels are increased as a consequence of the thyroidal adaptation to a PTU challenge.

Journal of Hypertension, 2004

The Journal of Comparative Neurology, 1991

This study examined the possibility that vasoactive intestinal peptide (VIP)-and substance P (SP)... more This study examined the possibility that vasoactive intestinal peptide (VIP)-and substance P (SP)-containing nerve fibers in bronchial smooth muscle, glands, epithelium, and blood vessels originate from neurons of airway ganglia. Explants of airway walls were maintained in culture with the expectation that nerve fibers from neurons of airway ganglia would remain viable, whereas fibers originating from neurons not present in the airway walls would degenerate. Airways were dissected and placed into culture dishes containing CMRL 1066 medium for 3,5, and 7 days. In controls (noncultured), VIP-and SP-like immunoreactivity was observed in nerve fibers associated with bronchial smooth muscle, glands, and blood vessel walls and in nerve cell bodies of airway ganglia. Nerve fibers containing SP were also observed within the bronchial epithelium. After 3,5, and 7 days in culture, VIP-and SP-containing fibers were identified in all of the same locations except in the airway epithelium where SP-containing fibers could not be demonstrated. VIP and SP were frequently colocalized in the same nerve fibers of bronchial smooth muscle and glands in controls and cultured airways. There were no statistically significant differences in nerve fiber density for either VIPor SP-containing fibers in bronchial smooth muscle between controlled and cultured airways. VIP concentrations in cultured airways were significantly less than in controls. The results suggest that a large proportion of VIP-and SP-containing nerve fibers supplying bronchial smooth muscle, glands, and blood vessels in the airways originate from neurons of airway ganglia.

Journal of Clinical Investigation, 2003

Previously we showed that neuropeptide Y (NPY), a sympathetic vasoconstrictor neurotransmitter, s... more Previously we showed that neuropeptide Y (NPY), a sympathetic vasoconstrictor neurotransmitter, stimulates endothelial cell migration, proliferation, and differentiation in vitro. Here, we report on NPY's actions, receptors, and mediators in ischemic angiogenesis. In rats, hindlimb ischemia stimulates sympathetic NPY release (attenuated by lumbar sympathectomy) and upregulates NPY-Y2 (Y2) receptor and a peptidase forming Y2/Y5-selective agonist. Exogenous NPY at physiological concentrations also induces Y5 receptor, stimulates neovascularization, and restores ischemic muscle blood flow and performance. NPY-mediated ischemic angiogenesis is not prevented by a selective Y1 receptor antagonist but is reduced in Y2-/mice. Nonischemic muscle vascularity is also lower in Y2-/mice, whereas it is increased in NPY-overexpressing rats compared with their WT controls. Ex vivo, NPYinduced aortic sprouting is markedly reduced in Y2-/aortas and spontaneous sprouting is severely impaired in NPY-/mice. NPY-mediated aortic sprouting, but not cell migration/proliferation, is blocked by an antifetal liver kinase 1 antibody and abolished in mice null for eNOS. Thus, NPY mediates neurogenic ischemic angiogenesis at physiological concentrations by activating Y2/Y5 receptors and eNOS, in part due to release of VEGF. NPY's effectiveness in revascularization and restoring function of ischemic tissue suggests its therapeutic potential in ischemic conditions.

Endocrinology, 1989

After subtotal thyroidectomy, the thyroid gland remnant undergoes compensatory alterations in fun... more After subtotal thyroidectomy, the thyroid gland remnant undergoes compensatory alterations in function and morphology. Under the trophic stimulation of elevated plasma TSH concentrations, the thyroid remnant responds with an increase in hormone synthesis and secretion and, in addition, increases in mass. We have examined the alterations in thyroid blood flow which accompany increased secretion and growth after hemithyroidectomy (HTX) in male Sprague-Dawley rats (200-220 g). At various times after surgical HTX (1, 2, and 3 weeks), blood samples for the determination of plasma hormone concentrations were obtained and tissue blood flows were determined using 15 +/- 5 microns diameter 141Ce-labeled microspheres in a modification of the reference sample microsphere technique. The microspheres were injected directly into the left cardiac ventricle via a 23-gauge needle passed through the chest wall while a reference blood sample was collected. After the animals were killed, tissues were cleaned and weighed, then tissue and reference blood sample radioactivities were determined. In addition, thyroidal immunoreactive vasoactive intestinal peptide was measured after acetic acid (0.67 N) extraction. After HTX, plasma TSH concentrations were significantly elevated. The plasma concentrations of T4 and T3 fell, but by less than the expected 50%. The mass of the remaining thyroid lobe increased progressively over the 3 weeks post thyroidectomy, reaching approximately 70% of the total thyroid gland weight of sham-operated controls. Thyroid blood flow per gram of tissue was significantly elevated at all times post HTX. HTX did not induce any alterations in thyroidal immunoreactive vasoactive intestinal peptide concentration. Thus, after HTX, the well documented compensatory alterations in thyroid remnant growth and secretion were accompanied by a prompt and striking increase in thyroid blood flow.

Endocrine Research, 1992

The presence of vasoactive intestinal peptide and neuropeptide Y in thyroid nerves and their effe... more The presence of vasoactive intestinal peptide and neuropeptide Y in thyroid nerves and their effects on thyroid blood flow are well known. However, the effects of these two neuropeptides on the various processes involved in thyroid hormone biosynthesis and release have not been fully explored. We have now tested these two peptides for effects on an early step in thyroid hormone biosynthesis, namely iodide uptake, a process which is comprised of trapping and organification. In these experiments, we have used anesthetized adult male rats pretreated with thyroxine or fed a low iodine diet to increase thyroidal sensitivity. Vasoactive intestinal peptide significantly increased iodide uptake in rats fed an iodine deficient diet but not in those fed a normal iodine diet. This effect disappeared if animals were pretreated with propylthiouracil. Neuropeptide Y did not alter iodide uptake in rats on either the low or the high iodine diet, regardless of the presence or absence of propylthiouracil. The effect of vasoactive intestinal peptide on iodide uptake could be due to its influence on the organification of iodine, or on thyroid blood flow, or on both processes.

Brain Research, 2005

In order to clarify the physiologic role of NPY in sensory processing, we obtained intracellular ... more In order to clarify the physiologic role of NPY in sensory processing, we obtained intracellular recordings of DRG neurons from wild type (WT) and NPY overexpressing transgenic rats (NPY-TG) before and after injury. We investigated medium and large diameter DRG neurons since upregulation of NPY peptide following the nerve injury occurs primarily in those cells. Neurons were classified as Aa/h and Ay using conduction velocity and action potential duration. Prior to the injury, Aa/h neurons of NPY-TG rats conducted more slowly and had a more brief AHP than similar cells from the WT group. Ay neurons at baseline conducted faster in TG animals compared to WT. Ligation of the 5th lumbar spinal nerve (SNL) produced certain changes in Aa/h cells that were evident only in the TG group. These include increased refractory period, increased input resistance, AHP prolongation and a depolarizing shift in threshold for AP initiation. The expected injury-induced CV slowing was not seen in NPY-TG Aa/h cells. In the Ay cell group, injury produced a depolarizing shift in the resting membrane potential, an increase in AP duration and decrease in AHP and refractory period duration only in WT rats, while NPY-TG cells lacked these injury-induced changes. Behavior tests showed diminished sensory response to nerve injury in NPY-TG rats, i.e., shorter duration of enhanced pain-related behavior and attenuation of contralateral effect. In conclusion, our observations suggest that NPY overexpression leads to reduced neuronal activity following nerve injury in a cell-specific manner.

American Journal of Physiology-Heart and Circulatory Physiology, 2007

A lentiviral construct for an enhanced green fluorescent protein (eGFP) driven by a chicken β-act... more A lentiviral construct for an enhanced green fluorescent protein (eGFP) driven by a chicken β-actin promoter, cytomegalovirus enhancer, and intronic sequences from rabbit β-globin (CAG) was used to produce transgenic lines of rats for evaluation of the usefulness of this approach in gene function studies. Fertilized eggs were collected from inbred Dahl S and outbred Sprague-Dawley rats, and ∼100 pl of concentrated virus were microinjected into the perivitrelline space of one-cell embryos. Of 121 embryos injected, 60 pups (49.6%) were born. Transgenic rates averaged 22% in Dahl S and 14% in Sprague-Dawley rats. Copy number ranged from one to four in the founders, and the inheritance of the transgene in a subsequent F1population was 48.2%. The small number of insertion sites enabled us to derive inbred transgenic lines with a single copy of the transgene within one generation. Sequencing of each transgene insertion site revealed that they inserted as single copies with a preference fo...