Kirk E. Dineley, Ph.D.

Associate Professor


Kirk Dineley, PhDMidwestern University
College of Osteopathic Medicine
Department of Pharmacology
Science Hall #322E
555 31st Street
Downers Grove, IL 60515

Office: (630) 960-3907
e-mail: kdineley@midwestern.edu

 

EDUCATION

BS Biology (Vertebrate Physiology), Pennsylvania State University
PhD Molecular Pharmacology, University of Pittsburgh

RESEARCH SUMMARY

The molecular mechanisms of steroidogenesis in the adrenal cortex

This research program is in collaboration with Drs. Latha Malaiyandi (Anatomy) and Phil Kopf (Pharmacology).

Excess secretion of the steroid hormone aldosterone from the adrenal cortex is associated with hypertension, stroke, myocardial infarction and other cardiovascular pathologies. At the cellular level, steroid production relies on complex signal transduction pathways, including alterations in cytoplasmic calcium. These signaling events regulate the activity of a network of steroidogenic enzymes, many of which are housed inside mitochondria. Our research relies mainly on a cell line (HAC15) derived from a human adrenocortical carcinoma to study calcium homeostasis and mitochondrial function using live-cell fluorescence microscopy techniques.

One project uses calcium-sensitive probes to evaluate cytoplasmic calcium changes that occur in response to various endogenous signaling molecules, including the hormones angiotensin and ACTH, and also neurotransmitters such as acetylcholine and ATP. Currently, we are interested in calcium oscillations that result from muscarinic receptor activation, and how that calcium activity may be related to the production and secretion of steroids. Using the calcium probe Fluo-4, we have shown that the muscarinic receptor agonists acetylcholine and carbachol cause intense oscillation of cytoplasmic calcium. These oscillations rely nearly completely on release and re-uptake of calcium stored in the endoplasmic reticulum. Antagonists that silence the oscillations suggest that M3 muscarinic receptors are the major subtype driving the oscillatory activity, whereas M2 receptors may have a smaller, inhibitory role that diminishes oscillations. We are currently working to determine the expression of different muscarinic receptors in HAC15 cells, and how the calcium oscillations influence the production and secretion of steroids.

Another focus of our research concerns the toxic effects of polybrominated diphenyl ethers (PBDEs) and other so-called "endocrine disruptors", which are thought to contribute to cardiovascular disease by increasing steroid production from the adrenal cortex. PBDEs are a class of flame retardants used in plastics, electronics, fabrics, and other common household materials. Although their industrial use is waning, PDBEs persist in the environment because they resist biodegradation. Concerns over their toxicity are further amplified by their lipophilic properties, which promote bioaccumulation, and penetration into the adrenal cortex, the womb, and breast milk. However, little is known regarding the long-term consequences of chronic PDBE exposure. Our preliminary observations show that the flame retardant BDE-47 stimulates steroid production in HAC15 cells and also in rats. At the cellular level, BDE-47 causes marked changes in cellular metabolism, and also alters the shape and number of mitochondria. Taken together, our results indicate that mitochondria may be an important intracellular target of flame retardants. In the larger picture, we anticipate that these studies will contribute importantly to our knowledge of cardiovascular disease particularly as it relates to environmental toxins.

Students interested in working with us should know that fluorescence methodologies and equipment are highly amenable to novice researchers, for a number of reasons: (i) the necessary devices are easy to operate, therefore researchers can obtain data after brief training periods; (ii) data generation is observed real-time, which allows the luxury and gratification of immediate experimental evaluation; and (iii) a vast array of available fluorescent probes permits the measurement of a large variety of cellular processes including ion fluxes, membrane potential, and mechanisms of cell death. We have mentored dozens of graduate and undergraduate students from all types of health sciences programs. Most have presented their work at regional or national meetings, and many are co-authors on posters and publications. Please contact us if you are interested.

Research projects

Project I: Intracellular calcium transients are mediated by activation of muscarinic receptors in adrenocortical carcinoma cells.

HAC15 cells demonstrate carbachol-induced transient intracellular free calcium flux, mediated primarily by activation of muscarinic receptors.

Project II: What is the role of mitochondria in flame retardant-induced hyperaldosteronism?

Using confocal microscopy, mitochondria (in green) and nuclei (in blue) are visualized using fluorophores. Exposure to the polybrominated flame retardant, BDE-47, produces mitochondrial morphology change in HAC15 cells. The movie represents a confocal z-stack of a BDE-47-treated HAC15 cell.

Selected Publications

Fluorescence detection of intracellular cadmium with Leadmium Green.

LM Malaiyandi, H Sharthiya, KE Dineley
Biometals. 2016 June; 29(4): 625-635

A comparison of Zn2+- and Ca2+-triggered depolarization of liver mitochondria reveals no evidence of Zn2+-induced permeability transition.
Devinney MJ, Malaiyandi LM, Vergun O, DeFranco DB, Hastings TG, Dineley KE
Cell Calcium. 2009 May; 45(5):447-55

The interaction of biological and noxious transition metals with the zinc probes FluoZin-3 and Newport Green.
Zhao J, Bertoglio BA, Devinney MJ Jr, Dineley KE, Kay AR
Anal Biochem. 2009 Jan 1; 384(1):34-41

Glutamate mobilizes Zn2+ through Ca2+-dependent reactive oxygen species accumulation.
Dineley KE
, Devinney MJ 2nd, Zeak JA, Rintoul GL, Reynolds IJ
J Neurochem. 2008 Sep; 106(5):2184-93

On the use of fluorescent probes to distinguish Ca2+ from Zn2+ in models of excitotoxicity.
Dineley KE
Cell Calcium. 2007 Sep; 42(3):341-2; author reply 343-4

Zinc causes loss of membrane potential and elevates reactive oxygen species in rat brain mitochondria.
Dineley KE
, Richards LL, Votyakova TV, Reynolds IJ
Mitochondrion. 2005 Feb; 5(1):55-65.

Direct visualization of mitochondrial zinc accumulation reveals uniporter-dependent and -independent transport mechanisms.
Malaiyandi LM, Vergun O, Dineley KE, Reynolds IJ
J Neurochem. 2005 Jun; 93(5):1242-50

Simultaneous detection of intracellular free calcium and zinc using fura-2FF and FluoZin-3.
Devinney MJ 2nd, Reynolds IJ, Dineley KE
Cell Calcium. 2005 Mar; 37(3):225-32

Divergent consequences arise from metallothionein overexpression in astrocytes: zinc buffering and oxidant-induced zinc release.
Malaiyandi LM, Dineley KE, Reynolds IJ
Glia. 2004 Mar; 45(4):346-53

Zinc inhibition of cellular energy production: implications for mitochondria and neurodegeneration.
Dineley KE
, Votyakova TV, Reynolds IJ
J Neurochem. 2003 May; 85(3):563-70.

Elevated intracellular zinc and altered proton homeostasis in forebrain neurons.
Dineley KE
, Brocard JB, Reynolds IJ
Neuroscience. 2002; 114(2):439-49

A reevaluation of neuronal zinc measurements: artifacts associated with high intracellular dye concentration.
Dineley KE
, Malaiyandi LM, Reynolds IJ
Mol Pharmacol. 2002 Sep; 62(3):618-27

The relationship between intracellular free iron and cell injury in cultured neurons, astrocytes, and oligodendrocytes.
Kress GJ, Dineley KE, Reynolds IJ
J Neurosci. 2002 Jul 15; 22(14):5848-55

Nitric oxide-induced changes in intracellular zinc homeostasis are mediated by metallothionein/thionein.
St Croix CM, Wasserloos KJ, Dineley KE, Reynolds IJ, Levitan ES, Pitt BR
Am J Physiol Lung Cell Mol Physiol. 2002 Feb; 282(2):L185-92

Vanilloid receptor expression suggests a sensory role for urinary bladder epithelial cells.
Birder LA, Kanai AJ, de Groat WC, Kiss S, Nealen ML, Burke NE, Dineley KE, Watkins S, Reynolds IJ, Caterina MJ
Proc Natl Acad Sci U S A. 2001 Nov 6; 98(23):13396-401

Induction of neuronal apoptosis by thiol oxidation: putative role of intracellular zinc release.
Aizenman E, Stout AK, Hartnett KA, Dineley KE, McLaughlin B, Reynolds IJ
J Neurochem. 2000 Nov; 75(5):1878-88

Astrocytes are more resistant than neurons to the cytotoxic effects of increased [Zn2+]i.
Dineley KE
, Scanlon JM, Kress GJ, Stout AK, Reynolds IJ
Neurobiol Dis. 2000 Aug; 7(4):310-20.

Involvement of RB kinases and phosphatases in life and death decisions.
An B, Dineley K, Zhang L, Termin T, Meijer L, Dou Q
Oncol Rep. 1997 Nov-Dec; 4(6):1129-34