The team used a common mouse model of heart failure to look for desmin clumps. In this model, the aorta -- the main artery coming from the heart -- is surgically constricted, which noticeably raises pressure and stress, and causes heart failure. After four weeks of pressure on the aorta, the mice develop symptoms of heart failure such as an enlarged heart and lung congestion. Desmin amyloid was more than doubled in the heart failure mice when using the same antibody and staining techniques used for the human tissue samples.

Then the researchers treated proteins from the mice hearts with epigallocatechin gallate (EGCG) -- a chemical from green tea known to break up amyloid. The treatment cut by half the amount of protein clumps.


"Interestingly, green tea has already been demonstrated to curb the incidence of cardiovascular disease as well as improve cognitive impairment in Alzheimer's models, though the mechanism for such action is unclear," says Agnetti. "EGCG's ability to 'de-clump' these sticky proteins could be one of green tea's healthy effects. Knowing how this chemical works could open new avenues for designing a new class of drugs that target protein clumping."

Next, the researchers wanted to identify the form of desmin that tended to clump. Based on their earlier work, they thought that one or more chemical phosphate groups added to the 27th or 31st building blocks in desmin's protein structure might affect how the protein clumps. They genetically engineered versions of desmin with one, both or none of the phosphate groups attached to desmin, tagged them with a green fluorescent signal to make them visible, and put them in heart cells using a virus.

A week later, using a microscope to track the green glow, the cells with desmin and two phosphate groups were still pumping, and this form of desmin was incorporated in the muscle fibers. The researchers say they believe this shows that the desmin with two phosphate groups is most likely the normal, healthy version of the protein.

The cells that had a single phosphate on desmin at the 31st position in the protein's chain of amino acids contracted more rapidly and had more green clumps, leading the researchers to believe that this behaves as the diseased version of the protein.

Agnetti learned from Richard O'Brien, M.D., Ph.D., a former Johns Hopkins neuroscientist now at Duke University, that PET is used to detect protein clumps in the brains of Alzheimer's and Parkinson's disease patients and can detect the clumps in certain genetic heart conditions that cause excessive protein clump formation. Following O'Brien's advice, the researchers tested if they could use this noninvasive technique to detect desmin clumps in mice with heart failure. Healthy and heart failure mice were injected with Amyvid, a radioactive dye that allows the researchers to see the protein clumps by PET. The heart failure mice had 13 percent more of the Amyvid taken up in their hearts than the healthy mice.

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