John P Konhilas
- Associate Professor
- Associate Professor, Cellular and Molecular Medicine
- Associate Professor, Molecular and Cellular Biology
- Associate Professor, Nutritional Sciences
- Acting Program Director
- Chair, Physiological Sciences - GIDP
- Associate Professor, Biomedical Engineering
- Gut microbiome and cardiac remodeling. Our lab has a long-standing interest in the ability of environmental factors, like diet, to impact cardiac disease. Advances in sequencing and bioinformatic technologies have allowed unprecedented characterization of the gut microbiome. We have discovered novel modifiers of the gut microbiome that protect against cardiac injury following ischemia.
- Sarcomere dynamics and crossbridge kinetics. Contractile perturbations downstream of Ca2+ binding to troponin C, the so-called sarcomere-controlled mechanisms, represent the earliest indicators of cardiovascular disease. We can now state the dynamics of cardiac contraction and relaxation during CVD are governed by downstream mechanisms, particularly the kinetics and energetics of the cross-bridge cycle. Our lab focuses on the contractile properties of the cardiomyocyte and how this changes with CVD.
- Sex dimorphisms in cardiac adaptation.Sex/gender differences exist in human cardiac disease resulting from many disease etoilogies including hypertension, myocardial infarction, and cardiomyopathies (HCM). We have adapted a novel model of menopause to uniquely address HCM and CVD, in general. As part of these studies, we became interested in a specific, energy-dependent signaling pathway, adenosine monophosphate-activated kinase (AMPK) demonstrating that AMPK regulates contractile function and energy cost of contraction.
- Predicting and mitigating postoperative surgical outcomes. (1) Cognitive impairment resulting from cardiac bypass surgery. Although treatment strategies for cardiovascular disease (CVD) are improving, coronary revascularization remains one of the most common interventional procedures. Following CABG surgery, cognitive impairment is reported in 50-75% of patients at discharge, 20-50% at 6 weeks and up to 40% at five years. Exciting new preclinical data from our group shows that systemic administration of Ang-(1-7) attenuates and even reverses CHF-induced cognitive impairment in mice. Our work has resulted in 2 patent applications (UA13-120 UA 14-167) and an IND application for the for Ang-(1-7) as a protective agent against CABG-induced cognitive impairment. (2) Predicting and mitigating postoperative new onset atrial fibrillation and cardiac remodeling. We have discovered a potential use for Human Amniotic Membranes for the prevention of postoperative (bypass surgery) outcomes. In human subjects, membrane placement during CABG preventative new onset postoperative atrial fibrillation. In mice, we prevented wall thinning post-myocardial infarction.
- Ph.D. Physiology and Biophysics
- University of Illinois at Chicago, Chicago, Illinois, United States
- The Molecular Mechanisms of Length-Dependent Activation in Striated Muscle
- M.S. Biological Sciences
- University of Illinois at Chicago, Chicago, Illinois, United States
- The Dynamics of Spherical Elastic Systems and its Significance to the Left Ventricular End-Systolic Pressure-Volume Relation
- B.A. Art and Art History
- Duke University, Durham, North Carolina, United States
- Certificate in Neurosciences Neuroscience
- Duke University, Durham, North Carolina, United States
No activities entered.
Cardio Muscle Bio & DiseasePSIO 484 (Spring 2018)
Cardio Muscle Bio & DiseasePSIO 584 (Spring 2018)
DissertationBME 920 (Spring 2018)
DissertationNSC 920 (Spring 2018)
Honors Independent StudyBIOC 499H (Spring 2018)
Honors ThesisPSIO 498H (Spring 2018)
Human PhysiologyPSIO 603A (Spring 2018)
Physiology SeriesPSIO 696A (Spring 2018)
Physiology Student ForumPS 696C (Spring 2018)
Physiology Student ForumPSIO 696C (Spring 2018)
Physiology/Biomed EngrBME 511 (Spring 2018)
Physiology/Biomed EngrPSIO 511 (Spring 2018)
ResearchNSC 900 (Spring 2018)
ResearchPSIO 900 (Spring 2018)
Rsrch Meth Biomed EngrBME 597G (Spring 2018)
Biology For Biomed EngrBME 510 (Fall 2017)
DissertationBME 920 (Fall 2017)
DissertationNSC 920 (Fall 2017)
Honors Independent StudyBIOC 499H (Fall 2017)
Honors Independent StudyMCB 499H (Fall 2017)
Honors ThesisPSIO 498H (Fall 2017)
Independent StudyMCB 399 (Fall 2017)
Physiology SeriesPSIO 696A (Fall 2017)
Physiology Student ForumPS 696C (Fall 2017)
Physiology Student ForumPSIO 696C (Fall 2017)
ResearchPSIO 900 (Fall 2017)
Research Methods In PsioPSIO 610 (Fall 2017)
Rsrch Meth Biomed EngrBME 597G (Fall 2017)
ThesisNSC 910 (Fall 2017)
Cardio Muscle Bio & DiseaseBME 484 (Spring 2017)
Cardio Muscle Bio & DiseaseBME 584 (Spring 2017)
Cardio Muscle Bio & DiseaseCMM 584 (Spring 2017)
Cardio Muscle Bio & DiseasePSIO 484 (Spring 2017)
Directed ResearchPSIO 492 (Spring 2017)
Honors Independent StudyPSIO 399H (Spring 2017)
Honors ThesisPSIO 498H (Spring 2017)
Human PhysiologyPSIO 603A (Spring 2017)
Independent StudyMCB 499 (Spring 2017)
Physiology/Biomed EngrBME 511 (Spring 2017)
ThesisNSC 910 (Spring 2017)
Biology For Biomed EngrBME 510 (Fall 2016)
Honors Independent StudyPSIO 399H (Fall 2016)
Honors ThesisPSIO 498H (Fall 2016)
Independent StudyMCB 299 (Fall 2016)
Independent StudyPSIO 399 (Fall 2016)
ThesisNSC 910 (Fall 2016)
- Lopez-Pier, M., Lipovka, Y., & Konhilas, J. P. (2016). Inherited Cardiomyopathies. In Cardiomyopathies. Intech Publishing.
- Schipper, D. A., Louis, A. V., Dicken, D. S., Johnson, K., Smolenski, R. T., Black, S. M., Runyan, R., Konhilas, J., Garcia, J. G., & Khalpey, Z. (2018). Improved metabolism and redox state with a novel preservation solution: implications for donor lungs after cardiac death (DCD). Pulmonary circulation, 7(2), 494-504.More infoLungs donated after cardiac death (DCD) are an underutilized resource for a dwindling donor lung transplant pool. Our study investigates the potential of a novel preservation solution, Somah, to better preserve statically stored DCD lungs, for an extended time period, when compared to low-potassium dextran solution (LPD). We hypothesize that Somah is a metabolically superior organ preservation solution for hypothermic statically stored porcine DCD lungs, possibly improving lung transplant outcomes. Porcine DCD lungs (n = 3 per group) were flushed with and submerged in cold preservation solution. The lungs were stored up to 12 h, and samples were taken from lung tissue and the preservation medium throughout. Metabolomic and redox potential were analyzed using high performance liquid chromatography, mass spectrometry, and RedoxSYS®, comparing substrate and pathway utilization in both preservation solutions. Glutathione reduction was seen in Somah but not in LPD during preservation. Carnitine, carnosine, and n-acetylcarnosine levels were elevated in the Somah medium compared with LPD throughout. Biopsies of Somah exposed lungs demonstrated similar trends after 2 h, up to 12 h. Adenosine gradually decreased in Somah medium over 12 h, but not in LPD. An inversely proportional increase in inosine was found in Somah. Higher oxidative stress levels were measured in LPD. Our study suggests suboptimal metabolic preservation in lungs stored in LPD. LPD had poor antioxidant potential, cytoprotection, and an insufficient redox potential. These findings may have immediate clinical implications for human organs; however, further investigation is needed to evaluate DCD lung preservation in Somah as a viable option for transplant.
- Avery, R. J., Yu, S. K., Cherukuri, G., Runyan, R. B., Konhilas, J., & Khalpey, Z. I. (2017). Remodeling Failing Human Myocardium With Hybrid Cell/Matrix and Transmyocardial Revascularization. ASAIO journal (American Society for Artificial Internal Organs : 1992).More infoGiven the limited treatment options for advanced heart failure, the intrinsic regenerative properties of stem cells have been evaluated for myocardial remodeling. Previous stem cells techniques for myocardiocyte remodeling have been limited by the low cellular retention. Presented is a hybrid approach for remodeling infarcted myocardium through implantation of allogeneic human amniotic fluid-derived mesenchymal stem cells within micronized human allograft-derived liquid matrix during the performance of transmyocardial revascularization (TMR). Given the induced increase in vascular density from TMR, we hypothesize that it may serve as a therapeutic delivery system for stem cell placement into damaged myocardium. We present a patient with ischemic cardiomyopathy and refractory angina, who clinically improved after this hybrid therapy of intraoperative TMR and placement of amniotic fluid-derived mesenchymal stem cells and liquid matrix within the TMR channels. Noninvasive testing of myocardial viability biomarkers utilizing both cardiac magnetic resonance imaging and thallium imaging supported the clinical improvement in cardiac symptom may be related to ventricular remodeling in a region of infarct with subsequent functional improvement.
- Danilo, C. A., Constantopoulos, E., McKee, L. A., Chen, H., Regan, J. A., Lipovka, Y., Lahtinen, S., Stenman, L. K., Nguyen, T. V., Doyle, K. P., Slepian, M. J., Khalpey, Z. I., & Konhilas, J. P. (2017). Bifidobacterium animalis subsp. lactis 420 mitigates the pathological impact of myocardial infarction in the mouse. Beneficial microbes, 8(2), 257-269.More infoThere is a growing appreciation that our microbial environment in the gut plays a critical role in the maintenance of health and the pathogenesis of disease. Probiotic, beneficial gut microbes, administration can directly attenuate cardiac injury and post-myocardial infarction (MI) remodelling, yet the mechanisms of cardioprotection are unknown. We hypothesised that administration of Bifidobacterium animalis subsp. lactis 420 (B420), a probiotic with known anti-inflammatory properties, to mice will mitigate the pathological impact of MI, and that anti-inflammatory T regulatory (Treg) immune cells are necessary to impart protection against MI as a result of B420 administration. Wild-type male mice were administered B420, saline or Lactobacillus salivarius 33 (Ls-33) by gavage daily for 14 or 35 days, and underwent ischemia/reperfusion (I/R). Pretreatment with B420 for 10 or 28 days attenuated cardiac injury from I/R and reduced levels of inflammatory markers. Depletion of Treg cells by administration of anti-CD25 monoclonal antibodies eliminated B420-mediated cardio-protection. Further cytokine analysis revealed a shift from a pro-inflammatory to an anti-inflammatory environment in the probiotic treated post-MI hearts compared to controls. To summarise, B420 administration mitigates the pathological impact of MI. Next, we show that Treg immune cells are necessary to mediate B420-mediated protection against MI. Finally, we identify putative cellular, epigenetic and/or post-translational mechanisms of B420-mediated protection against MI.
- Hay, M., Vanderah, T. W., Samareh-Jahani, F., Constantopoulos, E., Uprety, A. R., Barnes, C. A., & Konhilas, J. (2017). Cognitive impairment in heart failure: A protective role for angiotensin-(1-7). Behavioral neuroscience, 131(1), 99-114.More infoPatients with congestive heart failure (CHF) have increased hospital readmission rates and mortality if they are concomitantly diagnosed with cognitive decline and memory loss. Accordingly, we developed a preclinical model of CHF-induced cognitive impairment with the goal of developing novel protective therapies against CHF related cognitive decline. CHF was induced by ligation of the left coronary artery to instigate a myocardial infarction (MI). By 4- and 8-weeks post-MI, CHF mice had approximately a 50% and 70% decline in ejection fraction as measured by echocardiography. At both 4- and 8-weeks post-MI, spatial memory performance in CHF mice as tested using the Morris water task was significantly impaired as compared with sham. In addition, CHF mice had significantly worse performance on object recognition when compared with shams as measured by discrimination ratios during the novel object recognition NOR task. At 8-weeks post-MI, a subgroup of CHF mice were given Angiotensin (Ang)-(1-7) (50mcg/kg/hr) subcutaneously for 4 weeks. Following 3 weeks treatment with systemic Ang-(1-7), the CHF mice NOR discrimination ratios were similar to shams and significantly better than the performance of CHF mice treated with saline. Ang-(1-7) also improved spatial memory in CHF mice as compared with shams. Ang-(1-7) had no effect on cardiac function. Inflammatory biomarker studies from plasma revealed a pattern of neuroprotection that may underlie the observed improvements in cognition. These results demonstrate a preclinical mouse model of CHF that exhibits both spatial memory and object recognition dysfunction. Furthermore, this CHF-induced cognitive impairment is attenuated by treatment with systemic Ang-(1-7). (PsycINFO Database Record
- Kazui, T., Tran, P. L., Pilikian, T. R., Marsh, K. M., Runyan, R., Konhilas, J., Smith, R., & Khalpey, Z. I. (2017). A dual therapy of off-pump temporary left ventricular extracorporeal device and amniotic stem cell for cardiogenic shock. Journal of cardiothoracic surgery, 12(1), 80.More infoTemporary mechanical circulatory support device without sternotomy has been highly advocated for severe cardiogenic shock patient but little is known when coupled with amniotic stem cell therapy.
- Khalpey, Z. I., Runyan, R. B., Konhilas, J. P., & Kazui, T. (2017). A dual therapy of off-pump temporary left ventricular extracorporeal device and amniotic stem cell for cardiogenic shock. J Cardiothorac Surg, 7(12), 80.
- Luckey, S. W., Haines, C. D., Konhilas, J. P., Luczak, E. D., Messmer-Kratzsch, A., & Leinwand, L. A. (2017). Cyclin D2 is a critical mediator of exercise-induced cardiac hypertrophy. Experimental biology and medicine (Maywood, N.J.), 242(18), 1820-1830.More infoA number of signaling pathways underlying pathological cardiac hypertrophy have been identified. However, few studies have probed the functional significance of these signaling pathways in the context of exercise or physiological pathways. Exercise studies were performed on females from six different genetic mouse models that have been shown to exhibit alterations in pathological cardiac adaptation and hypertrophy. These include mice expressing constitutively active glycogen synthase kinase-3β (GSK-3βS9A), an inhibitor of CaMK II (AC3-I), both GSK-3βS9A and AC3-I (GSK-3βS9A/AC3-I), constitutively active Akt (myrAkt), mice deficient in MAPK/ERK kinase kinase-1 (MEKK1-/-), and mice deficient in cyclin D2 (cyclin D2-/-). Voluntary wheel running performance was similar to NTG littermates for five of the mouse lines. Exercise induced significant cardiac growth in all mouse models except the cyclin D2-/- mice. Cardiac function was not impacted in the cyclin D2-/- mice and studies using a phospho-antibody array identified six proteins with increased phosphorylation (greater than 150%) and nine proteins with decreased phosphorylation (greater than 33% decrease) in the hearts of exercised cyclin D2-/- mice compared to exercised NTG littermate controls. Our results demonstrate that unlike the other hypertrophic signaling molecules tested here, cyclin D2 is an important regulator of both pathologic and physiological hypertrophy. Impact statement This research is relevant as the hypertrophic signaling pathways tested here have only been characterized for their role in pathological hypertrophy, and not in the context of exercise or physiological hypertrophy. By using the same transgenic mouse lines utilized in previous studies, our findings provide a novel and important understanding for the role of these signaling pathways in physiological hypertrophy. We found that alterations in the signaling pathways tested here had no impact on exercise performance. Exercise induced cardiac growth in all of the transgenic mice except for the mice deficient in cyclin D2. In the cyclin D2 null mice, cardiac function was not impacted even though the hypertrophic response was blunted and a number of signaling pathways are differentially regulated by exercise. These data provide the field with an understanding that cyclin D2 is a key mediator of physiological hypertrophy.
- Vega, R. B., Konhilas, J. P., Kelly, D. P., & Leinwand, L. A. (2017). Molecular Mechanisms Underlying Cardiac Adaptation to Exercise. Cell metabolism, 25(5), 1012-1026.More infoExercise elicits coordinated multi-organ responses including skeletal muscle, vasculature, heart, and lung. In the short term, the output of the heart increases to meet the demand of strenuous exercise. Long-term exercise instigates remodeling of the heart including growth and adaptive molecular and cellular re-programming. Signaling pathways such as the insulin-like growth factor 1/PI3K/Akt pathway mediate many of these responses. Exercise-induced, or physiologic, cardiac growth contrasts with growth elicited by pathological stimuli such as hypertension. Comparing the molecular and cellular underpinnings of physiologic and pathologic cardiac growth has unveiled phenotype-specific signaling pathways and transcriptional regulatory programs. Studies suggest that exercise pathways likely antagonize pathological pathways, and exercise training is often recommended for patients with chronic stable heart failure or following myocardial infarction. Herein, we summarize the current understanding of the structural and functional cardiac responses to exercise as well as signaling pathways and downstream effector molecules responsible for these adaptations.
- Behunin, S. M., Lopez-Pier, M. A., Mayfield, R. M., Danilo, C. A., Lipovka, Y., Birch, C., Lehman, S., Tardiff, J. C., Gregorio, C. C., & Konhilas, J. P. (2016). Liver Kinase B1 complex acts as a novel modifier of myofilament function and localizes to the Z-disk in cardiac myocytes. Archives of biochemistry and biophysics.More infoContractile perturbations downstream of Ca(2+) binding to troponin C, the so-called sarcomere-controlled mechanisms, represent the earliest indicators of energy remodeling in the diseased heart . Central to cellular energy "sensing" is the adenosine monophosphate-activated kinase (AMPK) pathway, which is known to directly target myofilament proteins and alter contractility [2-6]. We previously showed that the upstream AMPK kinase, LKB1/MO25/STRAD, impacts myofilament function independently of AMPK . Therefore, we hypothesized that the LKB1 complex associated with myofilament proteins and that alterations in energy signaling modulated targeting or localization of the LKB1 complex to the myofilament. Using an integrated strategy of myofilament mechanics, immunoblot analysis, co-immunoprecipitation, mass spectroscopy, and immunofluorescence, we showed that 1) LKB1 and MO25 associated with myofibrillar proteins, 2) cellular energy stress re-distributed AMPK/LKB1 complex proteins within the sarcomere, and 3) the LKB1 complex localized to the Z-Disk and interacted with cytoskeletal and energy-regulating proteins, including vinculin and ATP Synthase (Complex V). These data represent a novel role for LKB1 complex proteins in myofilament function and myocellular "energy" sensing in the heart.
- Birch, C. L., Behunin, S. M., Lopez-Pier, M. A., Danilo, C., Lipovka, Y., Saripalli, C., Granzier, H., & Konhilas, J. P. (2016). Sex dimorphisms of crossbridge cycling kinetics in transgenic hypertrophic cardiomyopathy mice. American journal of physiology. Heart and circulatory physiology, 311(1), H125-36.More infoFamilial hypertrophic cardiomyopathy (HCM) is a disease of the sarcomere and may lead to hypertrophic, dilated, restrictive, and/or arrhythmogenic cardiomyopathy, congestive heart failure, or sudden cardiac death. We hypothesized that hearts from transgenic HCM mice harboring a mutant myosin heavy chain increase the energetic cost of contraction in a sex-specific manner. To do this, we assessed Ca(2+) sensitivity of tension and crossbridge kinetics in demembranated cardiac trabeculas from male and female wild-type (WT) and HCM hearts at an early time point (2 mo of age). We found a significant effect of sex on Ca(2+) sensitivity such that male, but not female, HCM mice displayed a decrease in Ca(2+) sensitivity compared with WT counterparts. The HCM transgene and sex significantly impacted the rate of force redevelopment by a rapid release-restretch protocol and tension cost by the ATPase-tension relationship. In each of these measures, HCM male trabeculas displayed a gain-of-function when compared with WT counterparts. In addition, cardiac remodeling measured by echocardiography, histology, morphometry, and posttranslational modifications demonstrated sex- and HCM-specific effects. In conclusion, female and male HCM mice display sex dimorphic crossbridge kinetics accompanied by sex- and HCM-dependent cardiac remodeling at the morphometric, histological, and cellular level.
- Keen, D. A., Constantopoulos, E., & Konhilas, J. P. (2016). The impact of post-exercise hydration with deep-ocean mineral water on rehydration and exercise performance. Journal of the International Society of Sports Nutrition, 13, 17.More infoDehydration caused by prolonged exercise impairs thermoregulation, endurance and exercise performance. Evidence from animal and human studies validates the potential of desalinated deep-ocean mineral water to positively impact physiological and pathophysiological conditions. Here, we hypothesize that deep-ocean mineral water drawn from a depth of 915 m off the Kona, HI coast enhances recovery of hydration and exercise performance following a dehydrating exercise protocol compared to mountain spring water and a carbohydrate-based sports drink.
- Lipovka, Y., & Konhilas, J. P. (2016). The complex nature of oestrogen signalling in breast cancer: enemy or ally?. Bioscience reports, 36(3).More infoThe pleiotropic nature of oestradiol, the main oestrogen found in women, has been well described in the literature. Oestradiol is positioned to play a unique role since it can respond to environmental, genetic and non-genetic cues to affect genetic expression and cellular signalling. In breast cancer, oestradiol signalling has a dual effect, promoting or inhibiting cancer growth. The potential impact of oestradiol on tumorigenesis depends on the molecular and cellular characteristics of the breast cancer cell. In this review, we provide a broad survey discussing the cellular and molecular consequences of oestrogen signalling in breast cancer. First, we review the structure of the classical oestrogen receptors and resultant transcriptional (genomic) and non-transcriptional (non-genomic) signalling. We then discuss the nature of oestradiol signalling in breast cancer including the specific receptors that initiate these signalling cascades as well as potential outcomes, such as cancer growth, proliferation and angiogenesis. Finally, we examine cellular and molecular mechanisms underlying the dimorphic effect of oestrogen signalling in breast cancer.
- Behunin, S. M., Lopez-Pier, M. A., Birch, C. L., McKee, L. A., Danilo, C., Khalpey, Z., & Konhilas, J. P. (2015). LKB1/Mo25/STRAD uniquely impacts sarcomeric contractile function and posttranslational modification. Biophysical journal, 108(6), 1484-94.More infoThe myocardium undergoes extensive metabolic and energetic remodeling during the progression of cardiac disease. Central to remodeling are changes in the adenine nucleotide pool. Fluctuations in these pools can activate AMP-activated protein kinase (AMPK), the central regulator of cellular energetics. Binding of AMP to AMPK not only allosterically activates AMPK but also promotes phosphorylation of AMPK by an upstream kinase complex, LKB1/Mo25/STRAD (liver kinase B 1, mouse protein 25, STE-related adaptor protein). AMPK phosphorylation by the LKB1 complex results in a substantial increase in AMPK activity. Molecular targeting by the LKB1 complex depends on subcellular localization and transcriptional expression. Yet, little is known about the ability of the LKB1 complex to modulate targeting of AMPK after activation. Accordingly, we hypothesized that differing stoichiometric ratios of LKB1 activator complex to AMPK would uniquely impact myofilament function. Demembranated rat cardiac trabeculae were incubated with varying ratios of the LKB1 complex to AMPK or the LKB1 complex alone. After incubation, we measured the Ca(2+) sensitivity of tension, rate constant for tension redevelopment, maximum tension generation, length-dependent activation, cooperativity, and sarcomeric protein phosphorylation status. We found that the Ca(2+) sensitivity of tension and cross-bridge dynamics were dependent on the LKB1 complex/AMPK ratio. We also found that the LKB1 complex desensitizes and suppresses myofilament function independently of AMPK. A phospho-proteomic analysis of myofilament proteins revealed site-specific changes in cardiac Troponin I (cTnI) phosphorylation, as well as a unique distribution of cTnI phosphospecies that were dependent on the LKB1 complex/ AMPK ratio. Fibers treated with the LKB1 complex alone did not alter cTnI phosphorylation or phosphospecies distribution. However, LKB1 complex treatment independent of AMPK increased phosphorylation of myosin-binding protein C. Therefore, we conclude that the LKB1/AMPK signaling axis is able to alter muscle function through multiple mechanisms.
- Khalpey, Z., Qu, N., Hemphill, C., Louis, A. V., Ferng, A. S., Son, T. G., Stavoe, K., Penick, K., Tran, P. L., Konhilas, J., Lagrand, D. S., & Garcia, J. G. (2015). Rapid porcine lung decellularization using a novel organ regenerative control acquisition bioreactor. ASAIO journal (American Society for Artificial Internal Organs : 1992), 61(1), 71-7.More infoTo regenerate discarded lungs that would not normally be used for transplant, ex vivo reseeding after decellularization may produce organs suitable for clinical transplantation and therefore close the donor gap. Organ regenerative control acquisition (Harvard Biosciences, Holliston, MA), a novel bioreactor system that simulates physiological conditions, was used to evaluate a method of rapid decellularization. Although most current decellularization methods are 24-72 hours, we hypothesized that perfusing porcine lungs with detergents at higher pressures for less time would yield comparable bioscaffolds suitable for future experimentation. Methods involved perfusion of 1% Triton X-100 (Triton) and 0.1% sodium dodecyl sulfate at varied physiological flow rates. Architecture of native and decellularized lungs was analyzed with hematoxylin and eosin (H&E) staining, transmission electron microscopy (TEM), and scanning electron microscopy (SEM). Dry gas and liquid ventilation techniques were introduced. Our 7 hour decellularization procedure removes nuclear material while maintaining architecture. Bioscaffolds have the microarchitecture for reseeding of stem cells. Hematoxylin and eosin staining suggested removal of nuclear material, whereas SEM and TEM imaging demonstrated total removal of cells with structural architecture preserved. This process can lead to clinical implementation, thereby increasing the availability of human lungs for transplantation.
- Konhilas, J. P., Chen, H., Luczak, E., McKee, L. A., Regan, J., Watson, P. A., Stauffer, B. L., Khalpey, Z. I., Mckinsey, T. A., Horn, T., LaFleur, B., & Leinwand, L. A. (2015). Diet and sex modify exercise and cardiac adaptation in the mouse. American journal of physiology. Heart and circulatory physiology, 308(2), H135-45.More infoThe heart adapts to exercise stimuli in a sex-dimorphic manner when mice are fed the traditional soy-based chow. Females undergo more voluntary exercise (4 wk) than males and exhibit more cardiac hypertrophy per kilometer run (18, 32). We have found that diet plays a critical role in cage wheel exercise and cardiac adaptation to the exercise stimulus in this sex dimorphism. Specifically, feeding male mice a casein-based, soy-free diet increases daily running distance over soy-fed counterparts to equal that of females. Moreover, casein-fed males have a greater capacity to increase their cardiac mass in response to exercise compared with soy-fed males. To further explore the biochemical mechanisms for these differences, we performed a candidate-based RT-PCR screen on genes previously implicated in diet- or exercise-based cardiac hypertrophy. Of the genes screened, many exhibit significant exercise, diet, or sex effects but only transforming growth factor-β1 shows a significant three-way interaction with no genes showing a two-way interaction. Finally, we show that the expression and activity of adenosine monophosphate-activated kinase-α2 and acetyl-CoA carboxylase is dependent on exercise, diet, and sex.
- Li, F., Buck, D., De Winter, J., Kolb, J., Meng, H., Birch, C., Slater, R., Escobar, Y. N., Smith, J. E., Yang, L., Konhilas, J., Lawlor, M. W., Ottenheijm, C., & Granzier, H. L. (2015). Nebulin deficiency in adult muscle causes sarcomere defects and muscle-type-dependent changes in trophicity: novel insights in nemaline myopathy. Human molecular genetics, 24(18), 5219-33.More infoNebulin is a giant filamentous protein that is coextensive with the actin filaments of the skeletal muscle sarcomere. Nebulin mutations are the main cause of nemaline myopathy (NEM), with typical adult patients having low expression of nebulin, yet the roles of nebulin in adult muscle remain poorly understood. To establish nebulin's functional roles in adult muscle, we studied a novel conditional nebulin KO (Neb cKO) mouse model in which nebulin deletion was driven by the muscle creatine kinase (MCK) promotor. Neb cKO mice are born with high nebulin levels in their skeletal muscles, but within weeks after birth nebulin expression rapidly falls to barely detectable levels Surprisingly, a large fraction of the mice survive to adulthood with low nebulin levels (
- Lipovka, Y., & Konhilas, J. P. (2015). AMP-Activated Protein Kinase Signalling in Cancer and Cardiac Hypertrophy. Cardiovascular pharmacology: open access, 4(3).More infoThe AMP-protein kinase (AMPK) pathway is very versatile as it regulates cellular energetic homeostasis in many different tissue types. An appreciation for the importance of AMPK signalling and regulation in cardiovascular and tumor biology is increasing. Recently, a link has been established between anti-cancer therapy and susceptibility to cardiac disease. It has been shown that some anti-cancer drugs lead to an increased risk of cardiac disease, underlined by de-regulation of AMPK signalling. This review explores the AMPK signalling axis in both cardiac and tumor metabolism. We then examine off-target AMPK inhibition by cancer drugs and how this may translate into increased risk of cardiovascular disease. Finally, we discuss the implication of deregulated AMPK signalling during different stages of cardiac hypertrophy. Better understanding of the molecular pathways behind pathological processes will lead to the development of more effective therapeutics for cancer and cardiovascular diseases.
- Lipovka, Y., Chen, H., Vagner, J., Price, T. J., Tsao, T., & Konhilas, J. P. (2015). Oestrogen receptors interact with the α-catalytic subunit of AMP-activated protein kinase. Bioscience reports, 35(5).More infoNormal and pathological stressors engage the AMP-activated protein kinase (AMPK) signalling axis to protect the cell from energetic pressures. Sex steroid hormones also play a critical role in energy metabolism and significantly modify pathological progression of cardiac disease, diabetes/obesity and cancer. AMPK is targeted by 17β-oestradiol (E2), the main circulating oestrogen, but the mechanism by which E2 activates AMPK is currently unknown. Using an oestrogen receptor α/β (ERα/β) positive (T47D) breast cancer cell line, we validated E2-dependent activation of AMPK that was mediated through ERα (not ERβ) by using three experimental strategies. A series of co-immunoprecipitation experiments showed that both ERs associated with AMPK in cancer and striated (skeletal and cardiac) muscle cells. We further demonstrated direct binding of ERs to the α-catalytic subunit of AMPK within the βγ-subunit-binding domain. Finally, both ERs interacted with the upstream liver kinase B 1 (LKB1) kinase complex, which is required for E2-dependent activation of AMPK. We conclude that E2 activates AMPK through ERα by direct interaction with the βγ-binding domain of AMPKα.
- Pollow, D. P., Romero-Aleshire, M. J., Sanchez, J. N., Konhilas, J. P., & Brooks, H. L. (2015). ANG II-induced hypertension in the VCD mouse model of menopause is prevented by estrogen replacement during perimenopause. American journal of physiology. Regulatory, integrative and comparative physiology, 309(12), R1546-52.More infoPremenopausal females are resistant to the development of hypertension, and this protection is lost after the onset of menopause, resulting in a sharp increase in disease onset and severity. However, it is unknown how a fluctuating ovarian hormone environment during the transition from perimenopause to menopause impacts the onset of hypertension, and whether interventions during perimenopause prevent disease onset after menopause. A gradual transition to menopause was induced by repeated daily injections of 4-vinylcyclohexene diepoxide (VCD). ANG II (800 ng·kg(-1)·min(-1)) was infused into perimenopausal and menopausal female mice for 14 days. A separate cohort of mice received 17β-estradiol replacement during perimenopause. ANG II infusion produced significantly higher mean arterial pressure (MAP) in menopausal vs. cycling females, and 17β-estradiol replacement prevented this increase. In contrast, MAP was not significantly different when ANG II was infused into perimenopausal and cycling females, suggesting that female resistance to ANG II-induced hypertension is intact during perimenopause. ANG II infusion caused a significant glomerular hypertrophy, and hypertrophy was not impacted by hormonal status. Expression levels of aquaporin-2 (AQP2), a collecting duct protein, have been suggested to reflect blood pressure. AQP2 protein expression was significantly downregulated in the renal cortex of the ANG II-infused menopause group, where blood pressure was increased. AQP2 expression levels were restored to control levels with 17β-estradiol replacement. This study indicates that the changing hormonal environment in the VCD model of menopause impacts the severity of ANG II-induced hypertension. These data highlight the utility of the ovary-intact VCD model of menopause as a clinically relevant model to investigate the physiological mechanisms of hypertension that occur in women during the transition into menopause.
- Chen, H., Perez, J. N., Constantopoulos, E., McKee, L., Regan, J., Hoyer, P. B., Brooks, H. L., & Konhilas, J. (2014). A method to study the impact of chemically-induced ovarian failure on exercise capacity and cardiac adaptation in mice. Journal of visualized experiments : JoVE.More infoThe risk of cardiovascular disease (CVD) increases in post-menopausal women, yet, the role of exercise, as a preventative measure for CVD risk in post-menopausal women has not been adequately studied. Accordingly, we investigated the impact of voluntary cage-wheel exercise and forced treadmill exercise on cardiac adaptation in menopausal mice. The most commonly used inducible model for mimicking menopause in women is the ovariectomized (OVX) rodent. However, the OVX model has a few dissimilarities from menopause in humans. In this study, we administered 4-vinylcyclohexene diepoxide (VCD) to female mice, which accelerates ovarian failure as an alternative menopause model to study the impact of exercise in menopausal mice. VCD selectively accelerates the loss of primary and primordial follicles resulting in an endocrine state that closely mimics the natural progression from pre- to peri- to post-menopause in humans. To determine the impact of exercise on exercise capacity and cardiac adaptation in VCD-treated female mice, two methods were used. First, we exposed a group of VCD-treated and untreated mice to a voluntary cage wheel. Second, we used forced treadmill exercise to determine exercise capacity in a separate group VCD-treated and untreated mice measured as a tolerance to exercise intensity and endurance.
- Khalpey, Z., Janardhanan, R., Konhilas, J., & Hemphill, C. (2014). First in man: adipose-derived stromal vascular fraction cells may promote restorative cardiac function. The American journal of medicine, 127(5), e11-2.
- Konhilas, J., Chen, H., Hwang, H., McKee, L. A., Perez, J. N., Regan, J. A., Constantopoulos, E., Lafleur, B., & Konhilas, J. -. (2013). Temporal and morphological impact of pressure overload in transgenic FHC mice. Frontiers in physiology, 4.More infoAlthough familial hypertrophic cardiomyopathy (FHC) is characterized as cardiac disease in the absence of overt stressors, disease penetrance, and pathological progression largely depend on modifying factors. Accordingly, pressure overload by transverse aortic constriction (TAC) was induced in 2-month-old, male mice with and without a FHC (R403Q) mutation in α-myosin heavy chain. A significantly greater number of FHC mice (n = 8) than wild-type (WT) mice (n = 5) died during the 9-week study period. TAC induced a significant increase in cardiac mass whether measured at 2 or 9 weeks post-TAC in both WT and FHC mice, albeit to a different extent. However, the temporal and morphological trajectory of ventricular remodeling was impacted by the FHC transgene. Both WT and FHC hearts responded to TAC with an early (2 weeks post-TAC) and significant augmentation of the relative wall thickness (RWT) indicative of concentric hypertrophy. By 9 weeks post-TAC, RWT decreased in WT hearts (eccentric hypertrophy) but remained elevated in FHC hearts. WT hearts following TAC demonstrated enhanced cardiac function as measured by the end-systolic pressure-volume relationship, pre-load recruitable stroke work (PRSW), and myocardial relaxation indicative of compensatory hypertrophy. Similarly, TAC induced differential histological and cellular remodeling; TAC reduced expression of the sarcoplasmic reticulum Ca(2+)-ATPase (2a) (SERCA2a; 2 and 9 weeks) and phospholamban (PLN; 2 weeks) but increased PLN phosphorylation (2 weeks) and β-myosin heavy chain (β-MyHC; 9 weeks) in WT hearts. FHC-TAC hearts showed increased β-MyHC (2 and 9 weeks) and a late (9 weeks) decrease in PLN expression concomitant with a significant increase in PLN phosphorylation. We conclude that FHC hearts respond to TAC induced pressure overload with increased premature death, severe concentric hypertrophy, and a differential ability to undergo morphological, functional, or cellular remodeling compared to WT hearts.
- Konhilas, J., McKee, L. A., Chen, H., Regan, J. A., Behunin, S. M., Walker, J. W., Walker, J. S., & Konhilas, J. -. (2013). Sexually dimorphic myofilament function and cardiac troponin I phosphospecies distribution in hypertrophic cardiomyopathy mice. Archives of biochemistry and biophysics, 535(1).More infoThe pathological progression of hypertrophic cardiomyopathy (HCM) is sexually dimorphic such that male HCM mice develop phenotypic indicators of cardiac disease well before female HCM mice. Here, we hypothesized that alterations in myofilament function underlies, in part, this sex dimorphism in HCM disease development. Firstly, 10-12month female HCM (harboring a mutant [R403Q] myosin heavy chain) mice presented with proportionately larger hearts than male HCM mice. Next, we determined Ca(2+)-sensitive tension development in demembranated cardiac trabeculae excised from 10-12month female and male HCM mice. Whereas HCM did not impact Ca(2+)-sensitive tension development in male trabeculae, female HCM trabeculae were more sensitive to Ca(2+) than wild-type (WT) counterparts and both WT and HCM males. We hypothesized that the underlying cause of this sex difference in Ca(2+)-sensitive tension development was due to changes in Ca(2+) handling and sarcomeric proteins, including expression of SR Ca(2+) ATPase (2a) (SERCA2a), β-myosin heavy chain (β-MyHC) and post-translational modifications of myofilament proteins. Female HCM hearts showed an elevation of SERCA2a and β-MyHC protein whereas male HCM hearts showed a similar elevation of β-MyHC protein but a reduced level of cardiac troponin T (cTnT) phosphorylation. We also measured the distribution of cardiac troponin I (cTnI) phosphospecies using phosphate-affinity SDS-PAGE. The distribution of cTnI phosphospecies depended on sex and HCM. In conclusion, female and male HCM mice display sex dimorphic myofilament function that is accompanied by a sex- and HCM-dependent distribution of sarcomeric proteins and cTnI phosphospecies.
- Konhilas, J., Perez, J. N., Chen, H., Regan, J. A., Emert, A., Constantopoulos, E., Lynn, M., & Konhilas, J. -. (2013). Effects of chemically induced ovarian failure on voluntary wheel-running exercise and cardiac adaptation in mice. Comparative medicine, 63(3).More infoThe role of exercise in decreasing the risk of cardiovascular disease in postmenopausal women has not been studied sufficiently. Accordingly, we investigated the effect of voluntary wheel-running and forced treadmill exercise on cardiac adaptation in mice treated with 4-vinylcyclohexine diepoxide (VCD), which selectively accelerates the loss of primary and primordial follicles and results in a state that closely mimics human menopause. Two-month-old female C57BL/6 mice injected with VCD (160 mg/kg) for 20 consecutive days underwent ovarian failure by 60 to 90 d after injection. Responses to voluntary wheel running and treadmill exercise did not differ between VCD- and vehicle-treated 7-mo-old C57BL/6 or outbred B6C3F1 mice. Moreover, adaptive cardiac hypertrophy, hypertrophic marker expression, and skeletal muscle characteristics after voluntary cage-wheel exercise did not differ between VCD- and vehicle-treated mice. Because 5' AMP-activated protein kinase (AMPK) is a key component for the maintenance of cardiac energy balance during exercise, we determined the effect of exercise and VCD-induced ovarian failure on the AMPK signaling axis in the heart. According to Western blotting, VCD treatment followed by voluntary cage-wheel exercise differently affected the upstream AMPK regulatory components AMPKα1 and AMPKα2. In addition, net downstream AMPK signaling was reduced after VCD treatment and exercise. Our data suggest that VCD did not affect exercise-induced cardiac hypertrophy but did alter cellular cardiac adaptation in a mouse model of menopause.
- Haines, C. D., Harvey, P. A., Luczak, E. D., Barthel, K. K., Konhilas, J. P., Watson, P. A., Stauffer, B. L., & Leinwand, L. A. (2012). Estrogenic compounds are not always cardioprotective and can be lethal in males with genetic heart disease. Endocrinology, 153(9), 4470-9.More infoHypertrophic cardiomyopathy (HCM) is more severe in male than female mice eating a soy-based diet. We sought to determine whether the detrimental effects are mediated by the phytoestrogens present in soy, the mechanism by which phytoestrogens act, and to test whether estrogen modulates the sexually dimorphic phenotype. A soy-free diet (casein based) supplemented with the predominant phytoestrogens in soy, genistein and daidzein, recapitulated the fibrotic, proapoptotic and negative hemodynamic effects of soy in male hearts. As with the soy diet, the hearts of female HCM mice were not negatively affected by the phytoestrogen-containing diet. To determine the role of estrogen in the sex differences mediated by diet in HCM, gonadectomies were performed and estrogen was administered to male and female HCM mice on a casein- or phytoestrogen-supplemented diet. Somewhat surprisingly, estrogen was not protective in male or female mice with HCM and, in fact, was lethal in phytoestrogen-fed male mice with HCM. Because genistein is a potent tyrosine kinase inhibitor and tyrosine kinase inhibition has been associated with cardiotoxicity, we tested its effects in isolated adult cardiac myocytes. Genistein inhibited different tyrosine kinases depending on sex and, in combination with estrogen, resulted in apoptosis only in adult male cardiac myocytes. Finally, we show that phytoestrogens led to distinct programs of gene expression in hearts from males vs. females with HCM, suggesting mechanisms by which males are more sensitive to the detrimental effects of phytoestrogens and females are protected. These results implicate the phytoestrogen genistein in mediating cardiac pathology in males with HCM and, importantly, establish that estrogen is not protective in the setting of HCM.
- Konhilas, J. P., Behunin, S. M., & Lynch, R. M. (2012). Keeping the beat. Focus on "enrichment of neonatal rat cardiomyocytes in primary culture facilitates long-term maintenance of contractility in vitro". American journal of physiology. Cell physiology, 303(12), C1218-9.
- Konhilas, J., Chen, H., Untiveros, G. M., McKee, L. A., Perez, J., Li, J., Antin, P. B., & Konhilas, J. -. (2012). Micro-RNA-195 and -451 regulate the LKB1/AMPK signaling axis by targeting MO25. PloS one, 7(7).More infoRecently, MicroRNAs (miR) and AMP-kinase (AMPK) have emerged as prominent players in the development of cardiac hypertrophy and heart failure. We hypothesized that components of the adenosine monophosphate-activated kinase (AMPK) pathway are targeted by miRs and alter AMPK signaling during pathological cardiac stress.
- Luczak, E. D., Barthel, K. K., Stauffer, B. L., Konhilas, J. P., Cheung, T. H., & Leinwand, L. A. (2011). Remodeling the cardiac transcriptional landscape with diet. Physiological genomics, 43(12), 772-80.More infoThe perception that soy food products and dietary supplements will have beneficial effects on cardiovascular health has led to a massive consumer market. However, we have previously noted that diet profoundly affects disease progression in a genetic model of hypertrophic cardiomyopathy (HCM). In this model, a soy-based diet negatively impacts cardiac function in male mice. Given the frequent connection between functional changes and transcriptional changes, we investigated the effect of diet (soy- vs. milk-based) on cardiac gene expression and how it is affected by the additional factors of sex and disease. We found that gene expression in the heart is altered more by diet than by sex or an inherited disease. We also found that the healthy male heart may be sensitized to dietary perturbations of gene expression in that it displays a gene expression profile more similar to diseased male and female hearts than to healthy female hearts. These observations may in part account for documented divergence in HCM phenotypes between males and females and between diets.
- Mark-Kappeler, C. J., Sen, N., Lukefahr, A., McKee, L., Sipes, I. G., Konhilas, J., & Hoyer, P. B. (2011). Inhibition of ovarian KIT phosphorylation by the ovotoxicant 4-vinylcyclohexene diepoxide in rats. Biology of reproduction, 85(4), 755-62.More infoIn vitro exposure of Postnatal Day 4 (PND4) rat ovaries to the occupational chemical 4-vinylcyclohexene diepoxide (VCD) destroys specifically primordial and primary follicles via acceleration of atresia. Because oocyte-expressed c-kit (KIT) plays a critical role in follicle survival and activation, a direct interaction of VCD with KIT as its mechanism of ovotoxicity was investigated. PND4 rat ovaries were cultured with and without VCD (30 μM) for 2 days. When assessed by Western analysis or mobility shift detection, phosphorylated KIT (pKIT) was decreased (P < 0.05) by VCD exposure, while total KIT protein was unaffected. Anti-mouse KIT2 (ACK2) antibody binds KIT and blocks its signaling pathways, whereas anti-mouse KIT 4 (ACK4) antibody binds KIT but does not block its activity. PND4 rat ovaries were incubated for 2 days with and without VCD with and without ACK2 (80 μg/ml) or ACK4 (80 μg/ml). ACK2 decreased pKIT; however, ACK4 had no effect. Conversely, ACK2 did not affect a VCD-induced decrease in pKIT, whereas ACK4 further reduced it. Because ACK2 and ACK4 (known to directly bind KIT) affect VCD responses, these results support the fact that VCD interacts directly with KIT. The effect of these antibodies on VCD-induced follicle loss was measured after 8 days of incubation. ACK2 further reduced (P < 0.05) VCD-induced follicle loss, whereas ACK4 did not affect it. These findings demonstrate that VCD induces ovotoxicity by direct inhibition of KIT autophosphorylation of the oocyte. The data also further support the vital function of KIT and its signaling pathway in primordial follicle survival and activation, as well as its role in VCD-induced ovotoxicity.
- Konhilas, J. P., Boucek, D. M., Horn, T. R., Johnson, G. L., & Leinwand, L. A. (2010). The role of MEKK1 in hypertrophic cardiomyopathy. International heart journal, 51(4), 277-84.More infoMEKK1 is a ubiquitously expressed mitogen activated protein kinase that is involved in tissue remodeling in a variety of settings including carotid artery blood flow cessation, wound healing, and breast adenocarcinoma intravasation. Here, we have tested the function of MEKK1 in genetic hypertrophic cardiomyopathy (HCM). MEKK1 was genetically deleted in C57Bl6/J mice expressing a mutant alpha-myosin heavy chain (HCM-MEKK1(-/-)). The absence of MEKK1 in HCM resulted in a more pronounced hypertrophy when compared to HCM mice with the MEKK1 gene intact without further increases in atrial natriuretic factor and beta-myosin heavy chain (MyHC) expression and fibrosis. Since MEKK1 is required for the induction of several tissue proteases, we tested the hypothesis that cardiac enlargement of HCM- MEKK1(-/-) mice was due to altered expression of urokinase-type plasminogen activator (uPA), JunB, matrix-metalloproteinase (MMP), and tissue inhibitors of MMPs (TIMPs). Because of its role in preventing apoptosis, we also tested the loss of MEKK1 on apoptotic mediators Bcl-2, cytochrome C, caspase-9, and caspase-3. uPA expression was decreased while JunB, MMP-9, caspase-9, and caspase-3 activities were elevated in HCM- MEKK1(-/-) hearts when compared to MEKK1(-/-), wild-type (WT), and HCM mice. Bcl-2 and Cyt C expression was elevated only in HCM mice. We conclude that the absence of MEKK1 induces a more pronounced cardiac hypertrophy to HCM through altered expression of proteases implicated in cardiac remodeling and increased apoptosis.
- Konhilas, J., & Konhilas, J. -. (2010). What we know and do not know about sex and cardiac disease. Journal of biomedicine & biotechnology, 2010.More infoCardiovascular disease (CVD) remains the single leading cause of death in both men and women. A large proportion of the population with CVD will die with a diagnosis of congestive heart failure (CHF). It is becoming increasingly recognized that sex differences exist in the etiology, development, and outcome of CHF. For example, compared to male counterparts, women that present with CHF are typically older and have systolic cardiac function that is not impaired. Despite a growing body of literature addressing the underlying mechanisms of sex dimorphisms in cardiac disease, there remain significant inconsistencies reported in these studies. Given that the development of CHF results from the complex integration of genetic and nongenetic cues, it is not surprising that the elucidation and subsequent identification of molecular mechanisms remains unclear. In this review, key aspects of sex differences in CVD and CHF will be highlighted with an emphasis on some of the unanswered questions regarding these differences. The contention is presented that it becomes critical to reference cellular mechanisms within the context of each sex to better understand these sex dimorphisms.
- Rosedale, R., Westman, E. C., & Konhilas, J. P. (2009). Clinical Experience of a Diet Designed to Reduce Aging. The journal of applied research, 9(4), 159-165.More infoOBJECTIVE: Aging is associated with elevated levels of glucose, insulin, and triglycerides. Our objective was to assess the effect of a nutritional program designed to reduce these correlates of aging. DESIGN: This is a retrospective chart review of patients attending an outpatient metabolic management program including a high-fat, adequate-protein, low-carbohydrate diet, nutritional supplementation and periodic individual visits. Outcomes measured at baseline and follow-up included body weight, fasting serum glucose, insulin, leptin, lipids, and thyroid hormone. RESULTS: Thirty-one patients were identified with complete information. The mean age of patients was 57.6 ± 2.4 consisting of 53% female and 47% male patients. The average duration between follow up visits was 91.5 ± 8.5 days. Of the parameters measured at the follow-up visit, body weight, serum leptin, insulin, fasting glucose, triglyceride, and free T(3) significantly decreased by 8.1 ± 0.8%, 48.2 ± 3.8%, 40.1 ± 4.7%, 7.6 ± 2.1%, 28.3 ± 5.7%, and 10.8 ± 1.8%, respectively. Furthermore, the triglyceride/high density lipoprotein ratio decreased from 5.1 ± 1.7 to 2.6 ± 0.5. CONCLUSIONS: In the context of an outpatient medical clinic, a high-fat, adequate-protein, low-carbohydrate diet with nutritional supplementation led to improvements in serum factors related to the aging process. Further research regarding this dietary approach and its relationship to aging is in order.