USF Health preclinical study finds that inhibiting lipoxygenase with a drug alters
innate inflammatory response, delaying heart tissue repair after cardiac injury

TAMPA, Fla. (May 10, 2021) — Blocking the fat-busting enzyme lipoxygenase with a synthetic inhibitor throws the immune system’s innate inflammatory response out of whack, compromising cardiac repair during acute heart failure, USF Health researchers found.

Their new preclinical study was published April 13 in Biomedicine & Pharmacotherapy.

In search of individualized heart failure therapies, Ganesh Halade, PhD, leads a USF Health Heart Institute team studying unresolved inflammation after heart attack. | Photo by Allison Long, USF Health Communications

Acute heart failure – triggered by a heart attack, severely irregular heartbeats, or other causes — occurs suddenly when the heart cannot pump enough blood to meet the body’s demands.

Following a heart attack or any cardiac injury, signals to immune cells called leukocytes carefully control physiological inflammation. Normally, there are two distinct but overlapping processes: an acute inflammatory response (“get in” signal), where leukocytes travel from the spleen to the injured heart to start removing dead or diseased cardiac tissue, and a resolving phase (“get out” signal), where inflammation is cleared with the help of macrophages that arrive to further repair the damage and form a stable scar.

A delay in either the initiation of inflammation or its timely clearance (resolution) can lead to impaired cardiac healing and progression to heart failure, said study principal investigator Ganesh Halade, PhD, an associate professor of cardiovascular sciences at the USF Health Morsani College of Medicine and a member of the USF Health Heart Institute.

The USF Health researchers applied three investigational approaches (in vitro, ex vivo, and in vivo) to assess whether a potent lipoxygenase (12/15 LOX) inhibitor ML351 could selectively alter inflammatory responses in adult mice following cardiac injury similar to a heart attack. Previous studies by Dr. Halade’s laboratory reported that lipoxygenase-deficient mice showed improved cardiac repair and heart failure survival after cardiac injury.

“We wondered if blocking a lipoxygenase with an external pharmacological compound (drug) would have the same beneficial effect — but the answer was no,” Dr. Halade said. “Instead, the collective results of our study indicate that ML351 dysregulated control of the normal physiological pathway of inflammation in cardiac repair, causing collateral damage.”

In the mice treated mice with ML351, leukocyte recruitment to the site of cardiac injury was delayed, which subsequently amplified inflammation at the site. At the same time, instead of leaving once the repair job was done, the immune cells remained at the site beyond the typical acute (and beneficial) inflammatory response phase. Basically, the late arrival (get-in signal) and delayed clearance (get-out signal) of immune cells impaired cardiac repair, Dr. Halade said.

A delay in either the initiation of inflammation or its timely clearance (resolution) can lead to impaired cardiac healing and progression to heart failure.

The latest study helps explain one more piece of the puzzle about the important role of immune-mediated acute inflammation and its clearance – both in promoting cardiac health and stopping the progression of heart failure, Dr. Halade said. Lipoxygenases, fatty-acid modifying enzymes that control metabolic and immune signaling, can promote either resolving (beneficial) or nonresolving (harmful) inflammation, he added.

“The take-home message is do not mess with (block) the lipoxygenase. Preserve it, because it’s a key enzyme for our defensive, innate immune response,” he said. “Knowing how drugs interact with the body’s precisely-balanced immune responses will be critical for understanding mechanisms to prevent, delay or treat the unresolved inflammation influencing heart failure.”

The USF Health study was supported by grants from the NIH’s National Heart, Lung and Blood Institute and the National Institute of Diabetes and Digestive and Kidney Diseases.

In search of individualized heart failure therapies, Ganesh Halade leads a USF Health Heart Institute team studying unresolved inflammation after heart attack

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Short-term inflammation is one of the body’s key defense mechanisms to help repair injury and fight infection. But low-level inflammation that does not subside has been linked to many common chronic conditions, including cardiovascular diseases such as atherosclerosis, atrial fibrillation and heart failure.

Ganesh Halade, PhD, an associate professor of cardiovascular sciences at the USF Health Morsani College of Medicine, investigates the safe clearance of acute inflammation – and what happens at the molecular and cellular levels when initially beneficial inflammation becomes harmful to the heart.  His team at the USF Health Heart Institute works on bridging the gap between the immune-responsive metabolism of fat and cardiac health by more clearly defining two distinct but simultaneous processes: the inflammatory response and how inflammation is safely cleared, or resolved.

In particular, Dr. Halade’s laboratory focuses on discovering ways to prevent, delay or treat the unresolved inflammation after a heart attack, which plays a key role in the pathology leading to heart failure. Their goal is to contribute to individualized therapies that may account for possible sex, racial/ethnic or age-related physiological differences in heart failure, a leading cause of hospitalizations and deaths worldwide.

Ganesh Halade, PhD, associate professor of cardiovascular sciences, joined the USF Health Heart Institute in February 2020. [Photo by Allison Long, USF Health Communications]

“Although several treatments and devices exist to help manage heart failure, the challenge remains the growth of metabolic risk factors like obesity, diabetes, hypertension and aging that amplify heart failure – and inflammation underlies all these conditions,” Dr. Halade said. “We’re in the early stages of understanding how the inflammatory response becomes chronic, or unresolved” after heart attack-induced injury.

Honing in on “the roots”

Dr. Halade’s late father, a farmer in Nashik close to Mumbai, India, emphasized to his young son that if he wanted to make a difference in life to “look to the roots, rather than the fruits.”

That philosophy drives Dr. Halade’s research endeavors. “We focus on the root causes of inflammation so that we can successfully treat the chronic inflammation that leads to heart failure,” he said.

Dr. Halade (center) with his research team, postdoctoral fellow Bochra Tourki, PhD, (left) and research associate Vasundhara Kain, PhD, (right). [Allison Long, USF Health Communications]

Clinical trials of several anti-inflammatory therapies so far have failed to show benefit in heart failure patients. Dr. Halade suggests that the investigational compounds intended to suppress inflammation very early in the cardiovascular disease process likely disrupt the tight control of immune-responsive signaling needed for timely resolution of inflammation.

“The inflammatory response and its resolution are two sides of the same coin – and they roll together. Blocking one side will affect the other,” he explained. “So, we don’t want to block the ‘get in signal’ needed to promote the early, ‘good’ inflammation. We want to accelerate the ‘get out’ signal to immune cells, so that as soon as repair of cardiac injury is done the acute inflammation leaves without becoming chronic.”

Dr. Halade views a high-resolution image (below) of a normally beating heart. [Photos by Anne DeLotto Baier, USFH Research Communications]

Connecting dysfunctional inflammation control and heart failure

A class of immune-system molecules orchestrates the resolution of tissue inflammation, an active process essential for advancing cardiac healing after a heart attack. These specialized proresolving mediators, or SPMs, are signaling molecules that form when fatty acids metabolize in response to immune activation of leukocytes.

Dr. Halade’s work is helping uncover new details on how heart failure-inducing inflammation may be limited (without promoting immunosuppression) – either by administering pharmacological SPMs, or activating enzymes that help stimulate the body’s own SPMs.

Over the last two years, he has published significant findings in several leading journals (papers summarized below) making the connections between fatty acids, inflammation control, and heart failure. Among Dr. Halade’s study collaborators is Charles Serhan, PhD, of Harvard Medical School, a pioneer in the emerging field of inflammation resolution.

• Science Signaling: This study followed the time course of inflammation and its resolution in a mouse heart attack model. The research showed for the first time that the active inflammation-resolving phase coincided with the acute inflammatory response facilitating cardiac repair after a heart attack. Among other factors, the researchers looked at types and amounts of SPMs, and the expression of enzymes that synthesize SPMs, both in the spleen and at the injured site of the heart. Macrophages, a type of white blood cell, are needed to generate SPMs as opposed to other immune cells, they reported.

Dr. Halade’s laboratory focuses on discovering ways to prevent, delay or treat the unresolved inflammation after a heart attack, which plays a key role in the pathology leading to heart failure. [Anne DeLotto Baier]

• Journal of the American Heart Association: The preclinical study discovered male-female cardiac repair differences in heart failure survival after heart attack, including improved recovery of cardiac function and greater survival of acute and chronic heart failure in female mice. Females generated higher levels of a particular fatty acid-derived signaling molecule (EET; epoxyeicosatrienoic acids) known to facilitate healing after a heart attack.

• ESC Heart Failure: The researchers profiled bioactive lipids (inflammatory biomarkers) in blood samples from hospitalized Black and White patients soon after a severe heart attack. They found a potent SPM signature (resolvin E1) was significantly lower in Black men and women than in Whites. The study concluded bioactive lipids are key for the diagnosis and treatment of cardiac repair after heart attack to delay heart failure.

• The FASEB Journal: Halade and colleagues identified a mouse model to study heart failure with preserved ejection fraction (HFpeF), a common form of heart failure linked to age-related obesity. Using this unique model of obese aging, they defined how the deficiency of a single resolution receptor triggers obesity in mice at an early age, which can give rise to many of the molecular and cellular processes ultimately leading to HFpEF.

Vasundhara Kain (seated) and Bochra Tourki, look at slides for a paper on age-related obesity and heart failure. [Allison Long, USF Health Communications]

Insight into potential inflammation-resolving therapies

As they learn more about the metabolic and immune-responsive signals that control acute cardiac inflammation, researchers hope to harness the capacity of fatty acid-derived bioactive molecules to prevent, diagnose and treat heart failure, Dr. Halade said. SPMs are derived primarily from omega-3 fats in our diet – the polyunsaturated “good” fats in foods like salmon, avocados, almonds, and walnuts.

Some evidence indicates that omega 3-rich diets and/or SPM supplements, as well as getting enough exercise and quality sleep may help prevent the unresolved inflammation leading to heart failure, Dr. Halade said. If SPMs are not produced due to risk factors like obesity or aging, or because enzymes required to metabolize fatty acids are deficient, then drugs specifically designed to facilitate cardiac repair and calm inflammation might delay or treat heart failure, he added. Distinctive biochemical signatures acquired by analyzing SPMs or other metabolites might even be used to help diagnose heart failure or predict which treatments will work best for certain patients.

Dr. Halade joined USF Health this February from the University of Alabama at Birmingham, where he was a faculty member since 2013. He received his PhD in pharmacology from the University of Mumbai Institute of Chemical Technology in 2007. He completed two postdoctoral fellowships at the University of Texas Health Science Center in San Antonio. The first fellowship focused on nutritional immunology. The second was conducted with mentor Merry Lindsey, PhD, to examine the effects of obesity on post-heart attack cardiac structure and function.

Foods rich in omega-3 fatty acids (including salmon, walnuts and avocados), as well as enough exercise and quality sleep, may help prevent unresolved inflammation contributing to cardiovascular disease.

Dr. Halade’s research is supported by funding from the NIH’s National Heart, Lung and Blood Institute. In 2018, he received American Physiological Society Research Career Enhancement Award to train in lipidomics at the RIKEN Center for Integrative Medical Sciences in Japan.

His inflammation resolution research has been recognized with two awards for studies published in the American Journal of Physiology-Heart and Circulatory. An Article Impact Award 2020 was conferred this March by the American Physiological Society for Dr. Halade’s work defining the impact of the cancer drug doxorubicin on the heart and spleen. He also received a 2017 Best Paper Award from the Unbound Science Foundation. Dr. Halade is associate editor for the American Journal of Physiology-Heart and Circulatory and for Scientific Reports, and serves on the editorial boards of several other high-impact journals in cardiovascular sciences.

At left: Beneficial resolution of inflammation following cardiac repair. At right: Risk factors like aging, obesity and some medications can contribute to unresolved (chronic) inflammation, which impairs cardiac repair and can lead to heart failure. [Graphic courtesy of Ganesh Halade]

Some things you may not know about Dr. Halade

• As an undergraduate student in India, Dr. Halade won the gold medal in fencing at a statewide collegiate competition.
• To help promote a heart healthy lifestyle, he enjoys recreational bicycling and gardening in his backyard, where he grows vegetables and chiles.
• Halade lives in Tampa with his wife Dipti, an information technology engineer, and their son Arav, 13.

Top:  Sources of inflammation include injury (like damage from a heart attack), infection (viruses, bacteria or other pathogens), and factors associated with lifestyle (such as poor diet and lack of exercise). Below: Ways to help prevent unresolved cardiac inflammation associated with lifestyle. [Graphics courtesy of Ganesh Halade]

-Video by Allison Long, USF Health Communications and Marketing