Chapter Seventeen: Problematic MH Research: Muscle Calcium Uptake

Chapter 17 Sub-sections

An article on a case of human MH and its successful treatment via cardiac bypass began a sad era in testing a person for susceptibility (Ryan et al, 1974). This paper proposed measuring calcium uptake from muscle strips as a test for detecting MH susceptibility (Mabuchi et al, 1978). However there was a major contradiction to its use in MH testing: the test was positive immediately after the MH episode while the patient was still recovering, and normal later, when the patient had recovered more fully. Little did we know that this application was the beginning of a wayward path into MH testing.

Muscle calcium uptake did not have publications supporting its application in MH. MH experts became aware of calcium uptake as a test primarily via diagnostic letters to patients, signed by John Ryan, an anesthesiologist at Harvard's Massachusetts General Hospital. Ryan et al initially reported cardiopulmonary bypass as a treatment of acute MH (Ryan et al, 1974). As mentioned above, the problem was that calcium uptake had been abnormal immediately after the MH episode, but normal in the same patient several months later, i.e., poor evidence for reliability.

Letters to patients (personal files) who had been tested by calcium uptake did not include evidence supporting its validity. Their impact could not be ignored, as the test came from the muscle research laboratory of Harvard University. It utilized thin strips of muscle from a small biopsy that were frozen and shipped to Boston. It was easier and more convenient than the contracture test, which utilized viable twitching muscle specimens and therefore required the patient to travel to the test center for careful dissection during surgery and immediate delivery to the muscle laboratory.

The Harvard laboratory had several publications concerning muscle calcium uptake, but none directly supportive of its use as an MH diagnostic test (Sreter et al, 1964, 1966; Sreter, 1969; Mabuchi et al, 1978). Many involved in MH doubted the efficacy of the calcium uptake test. It is an in vitro measure of total calcium bound to muscle tissue, not isolated portions. It is not as sensitive as calcium binding, a measure of calcium taken up into isolated SR, and it did not distinguish normal humans from MH susceptible humans (Blanck et al, 1981). Next was an abortive attempt to compare contracture testing and muscle calcium uptake.

Initial Comparative Contracture-calcium Uptake Study, Unilaterally Cancelled

In an effort to document the accuracy of muscle calcium uptake, Henry Rosenberg, then of Philadelphia, began a joint study with the Harvard laboratory to compare MH patient contracture test results with those of calcium uptake, in the same patients. Dr. Rosenberg's laboratory performed contracture studies on a patient sample and forwarded an unused portion of that sample to Dr. Ryan's laboratory for study of calcium uptake. When six patients had been studied, the results disagreed, and Ryan, despite Rosenberg's protests, unilaterally discontinued the study. Ryan believed that the test was accurate despite the conflicting contracture results.

This vexing decision by Ryan was disturbing and I discussed this with my chair at Mayo, and with the chair at Harvard in fall 1983, at the annual meeting in Atlanta of the American Society of Anesthesiologists, and the oral examinations of the American Board of Anesthesiology in Albuquerque. Both supported a cooperative study between their two institutions, that would be blinded, and that would be finished.

Harvard/Mayo Contracture-calcium Uptake Study that could Not be Cancelled

The two chairs, Ryan, and I planned and approved a double blind study similar to that of Rosenberg and Ryan, in which no one would know results until the study was completed. Furthermore, an outside academic, with no knowledge of the study until requested to become involved, would supervise breaking the code. After the study was completed, the chair of anesthesia at Columbia University, agreed to break the code, and in a letter confirmed his role (personal files).

The study went smoothly. We biopsied thigh muscle on patients with questions regarding susceptibility to MH. My Mayo laboratory measured muscle contractures, and saved a portion for shipment to Ryan, both sets of samples coded as to identity. He didn't know our contracture results, and we didn't know his calcium uptake results. When eight or ten samples had accumulated, a technician blinded to the code shipped these to Harvard for muscle calcium uptake.

When finished, we had 29 joint samples, 28 suitable for publication. No results were known until we broke the code in a conference telephone call with the anesthesia chair at Columbia. The results did not support the accuracy of muscle calcium uptake, which had false positive results. Ryan was disturbed that conflicting results had again occurred, this time in a completed protocol. He and I began to prepare a manuscript for publication. After several unproductive efforts to consolidate our differences in writing the manuscript, I submitted our individual efforts to a journal, with a letter explaining the problems, and requested the aid of the editor-in-chief.

The editor was supportive and attempted to consolidate our differences, but was not successful. He described Ryan's draft as unsuitable for a peer-review journal. I knew now that I needed to consult someone for advice who had no knowledge of the situation, and phoned my friend Arthur Keats, now deceased, a former editor-in-chief of the journal, Anesthesiology, who was noted for his common sense and straightforward approach. He had tutored me as a junior examiner on the anesthesia boards, and I knew I could trust in an objective opinion that favored no one. He said that this was a sad story, but, when you lose a colleague, you lose a colleague's data, so forget it. Arthur had quite a formidable presence but a subtle sense of humor and used it in the board examination system. I provide an example later in this memoir when describing the American Board oral examination.

A Direct Contact from Harvard

During our efforts on the calcium uptake manuscript, the late senior Harvard Anesthesia Professor, Aaron Gissen, phoned. He was chair of an ad hoc Harvard committee examining the validity of the muscle calcium uptake test. Controversies concerning the test had prompted the university to establish his committee. Gissen informed me that a professor from another institution (personal files), a former President of the American Society of Anesthesiologists, had advised him to contact me. Gissen said that two members on the ad hoc committee (identities, personal files) were involved with the calcium uptake project, but that neither had mentioned the Mayo-Harvard study. He learned of it only after talking to me, during the several conversations in which we had phoned each other, and in which I described the study and our results.

Once the entire project was finished, in part because of the Ryan-Rosenberg brouhaha, I reviewed my records of letters, phone calls, and other related sources. My files relating to this specific study include more than 65 letters, phone calls, and other material involving members of both anesthesia departments, present and former officers of the American Society of Anesthesiologists, and others. Some who contacted me were concerned with success in publication, some with a major public relations problem for our specialty, and others with legal ramifications, institutional and personal. Those active in MH testing were vitally interested in this study and were disturbed to see its publication fail, particularly since it duplicated the results of the Rosenberg-Ryan study.

In June 1986, the Harvard investigators promoting calcium uptake testing published an article describing results in 121 patients, 76 of whom had survived unequivocal episodes as diagnosed by standard criteria (Allen et al, 1986).

They concluded: "… the decreased SR Ca2+ uptake correlates so well with MH that it clearly may provide a valuable diagnostic tool."

But the test involves thin strips of muscle, not isolated SR, and they provided no details justifying these 76 ‘unequivocally' susceptible patients. This appears to be enthusiasm untempered by discipline, and in part led to calcium uptake being applied in determining care of patients (Schwartz et al, 1984; Watson et al, 1986), although that was strongly criticized (Ellis et al, 1986).

Unexpected: Harvard Abandons the Calcium Uptake Test

Then, a surprise: Harvard abandoned the test in late 1986. The Pathology Department representative commented in the MHAUS (Malignant Hyperthermia Association of the United States) Communicator that they were abandoning the calcium uptake test (spring and fall issues, 1987). They had performed the assay since fall 1985, and could not discriminate between normal and abnormal patients.

In the spring issue, Ryan stated that he was at a loss concerning the results of the Pathology Department, and that his group had successfully distinguished between patients with positive histories and normal controls for 12 years. They had examined more than 1200 muscle specimens and counseled more than 600 families. Laboratories could duplicate contracture test results, but not those of calcium uptake.

Verification of the Failure of the Calcium Uptake Test

In April, 1987, Muldoon's laboratory in the Department of Anesthesiology at the Uniformed Services University of the Health Sciences published a comprehensive refutation of the calcium uptake test (Nagarajan et al, 1987). It had performed contracture tests for diagnosis in 51 patients in the preceding years, and had frozen extra tissue. Her colleagues had now performed calcium uptake measurements on the preserved muscle tissue. Calcium uptake values had not declined in specimens stored up to 5 years at -70o C, -94o F. There was no correlation with any accepted prognosticators of MH susceptibility. They stated that calcium uptake data also may fail to correlate with results from isolated fragments of sarcoplasmic reticulum. Finally, re-biopsy of a patient originally tested by calcium uptake confirmed its unreliability (Jaffe et al, 1991). Experts in MH could now focus comfortably on promising tests.