Dysregulated MicroRNAs Linked to Muscular Disorders

HMS researchers have analyzed the regulatory involvement of over 400 microRNAs (miRNAs) associated with muscular disorders in an effort to shed light on the pathogenesis of these conditions.

Courtesy Iris Eisenberg

Disease patterns. Principal component analysis (PCA) mapping shows differences in expression of microRNAs in Duchene muscular dystrophy (DMD) and normal muscle. PC1 accounts for 28.7 percent of the variance in the dataset and includes noise such as different muscle type and gender. PC2 accounts for 15.1 percent of the variance and separates DMD (dark green) from normal muscle (light green). Becker muscular dystrophy (BMD, in red), the milder cousin of DMD, appears between the DMD and normal muscle. Severe cases of BMD were closer to DMD, and milder forms were nearer the normal muscle. (The labels are patient reference numbers.)

Their microarray research implicated 185 dysregulated miRNAs in the development of 10 muscular disorders in humans, as reported in the Oct. 23 Proceedings of the National Academy of Sciences. Senior author Louis Kunkel, HMS professor of pediatrics at Children’s Hospital Boston, and lead author Iris Eisenberg, an HMS research fellow in pediatrics, compared miRNA expression patterns in 80 muscle specimens from patients with muscular dystrophies and myopathies.

Of the 185 miRNAs, a total of 151 were upregulated, 28 were downregulated, and six were expressed in an inconsistent pattern across disorders. Five were dysregulated in all samples and may provide clues about an underlying regulatory pathway in these disorders.

When the single-stranded rogue miRNAs were either up- or down-regulated, they interfered with messenger RNA expression and acted as negative regulators, with an inverse correlation between miRNA expression and gene expression.
Duchenne muscular dystrophy (DMD) and Miyoshi myopathy had the strongest functional correlations with this mRNA and miRNA seesaw effect. The researchers identified 57 mRNA and miRNA interactions in DMD.

Some of the muscle disorders were associated with exclusively dysregulated miRNAs, pointing to a distinctive regulatory mechanism in particular cases. DMD—whose mutant gene Kunkel was the first to isolate, in 1986—had 62 dysregulated miRNAs, 14 that were solely dysregulated in this phenotype. Other uniquely dysregulated miRNAs showed up in disorders like limb girdle muscular dystrophy and facioscapulohumeral muscular dystrophy (FSHD), suggesting a unique role of miRNAs in the pathology of muscular disorders. A subgroup of 18 dysregulated miRNAs gave researchers enough evidence to distinguish between specific diseases and controls.

Some unexpected miRNAs also cropped up in the microarrays. Eleven previously identified as related to immunity were dysregulated. “In normal muscle, you don’t see any of these inflammatory cells,” Kunkel said. The findings were consistent with the observation of elevated levels of immune cells and inflammatory cytokines in the extracellular environment around dystrophic muscle.

Eisenberg hopes that miRNAs will give researchers alternative therapeutic targets. Two promising therapies for muscular disorders exist: one replaces the defective gene with a normal one using virus vectors; the other induces exon skipping of a mutation-containing exon to produce a frame shift and a smaller but functional protein. Beyond these, there is only palliative and symptomatic relief for patients.
Said Kunkel, “If you had a microRNA that targeted very specific mRNA, its up- or down-regulation could influence their gene expression; you could influence a whole pathway by just targeting one microRNA.”

—Laura Geggel

What Med Students Don’t Know About the ‘Doctor Draft’

Medical students are ill informed about military medical ethics, the Geneva Conventions, and laws governing the drafting of doctors, according to a study from the Cambridge Health Alliance.

The Internet-based questionnaire surveyed 1,756 medical students at eight medical schools across the country. Of those, about 94 percent replied that they had received less than one hour of instruction about military medical ethics and only 3.5 percent were familiar with legislation regarding the “doctor draft.” In 1987, Congress approved the draft plan, called the Health Care Personnel Delivery System (HCPDS), which gives the legislative and executive branches the power to begin drafting civilian physicians within a matter of weeks of implementing the authorized HCPDS plan.

“The HCPDS allows few exemptions and assumes a priori that physicians practicing in the civilian sector are physically fit for military service,” the study reports; it appears in the current quarterly issue of International Journal of Health Services.

Nearly 34 percent of respondents were unaware that the Geneva Conventions mandate that physicians “treat the sickest first, regardless of nationality,” and the same percentage could not state when it would be appropriate to disobey an unethical order.

“Medical students should be taught more about respect for human rights in general and military medical ethics in particular,” said Wesley Boyd, lead author of the study and an HMS clinical instructor in psychiatry at Cambridge Hospital. Steffie Woolhandler, HMS associate professor of medicine at Cambridge Hospital and Massachusetts General Hospital, is the senior author.

Approximately five percent of those surveyed had past, current, or future military obligations. Of this group, about 15 percent were aware of HCPDS. More of those with military obligations than those without said they were very familiar with the Geneva Conventions. Yet their answers revealed they were no more informed than their classmates.

“One surprising finding is that over six percent of our respondents said they would be willing to kill a detainee with a lethal injection and would not countermand an order to do so,” Boyd said.

Responses did not vary significantly by gender, expected year of graduation, expected choice of medical specialty, or medical school.

—Laura Geggel

Diet Modifications May Counter Infertility

HSPH scientists have found that women who follow a combination of at least five low-risk lifestyle factors, including diet, weight control, and physical activity, can decrease their risk of ovulatory-disorder infertility by 69 percent compared to women who follow between zero and four low-risk lifestyle factors.

The nutritionists first focused on polycystic ovary syndrome (PCOS), a condition that contributes to several disorders, including menstrual irregularity and infertility. PCOS appears to be linked to insulin resistance, which in turn can be influenced by diet.

“We wanted to know whether anything influencing insulin sensitivity, including diet, would also improve reproductive function in otherwise healthy women and not only women with PCOS,” said the study’s lead author, Jorge Chavarro, a research fellow in the Department of Nutrition at HSPH. Walter Willett, chair of the department and the Fredrick John Stare professor of epidemiology and nutrition, is the senior author.

For every six couples who try to get pregnant, one pair faces infertility; ovulatory disorders affect 18 to 30 percent of these couples. To identify a fertility diet, the HSPH researchers followed 17,544 women without a history of infertility who were trying to or became pregnant during the eight-year prospective study, which appears in the Nov. 1 issue of Obstetrics & Gynecology.

Women who had the highest fertility score consumed less trans fat, more vegetable protein, more carbohydrates with high fiber and a low glycemic index, more nonheme iron, more multivitamins, and more high-fat dairy products, among other choices.

The high-fat dairy product component may be surprising. Galactose studies in mice, for example, have linked the sugar to premature ovarian failure. And a 1994 study from Brigham and Women’s Hospital reported that ingesting galactose may negatively affect ovarian function.

Yet, said Chavarro, “we found that low-fat dairy products were, in fact, associated with a higher risk of infertility.” He is unsure why this is the case but speculated that bovine hormones may be present in higher quantities in high-fat milk and could facilitate women’s fertility.

In addition to diet, the study analyzed body mass index (BMI). The researchers graphed a U-shaped relationship between BMI and ovulatory- disorder infertility; women with a BMI between 20 and 25 had a lowered infertility risk while women with a BMI on either side of this range had a similar heightened risk of infertility.

Vigorous exercise—more than 30 minutes per day—had no effect on ovulatory disorders until the researchers removed BMI from the equation, showing that exercise led to a slightly lower risk for ovulatory infertility.

The study found that fertility rose for each additional low-risk lifestyle factor a woman followed. Women who abided by five or more factors experienced a sixfold decrease in risk compared to women who followed none.

—Laura Geggel

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