A critical value for total calcium was reported on a 68-year-old female patient who was admitted to the emergency department 15 h prior. The patient's laboratory results were significant for a serum total calcium of 16.8 (8.4–10.3) mg/dL, which was verified by replicate analysis, and her free calcium concentration of 2.25 (1.12–1.32) mmol/L. The patient's serum phosphate was low at 0.5 (2.7–4.5) mg/dL, and her parathyroid hormone (PTH)2 was also suppressed at 6 (15–65) pg/mL. A review of the patient's medical records revealed a past medical history significant for hypertension, hyperlipidemia, and non–insulin-dependent diabetes mellitus. Her medication list included aspirin (81 mg daily), pravastatin (80 mg daily), and verapamil (120 mg daily).
Calcium exists in 3 states: approximately 50% ionized, 40% protein-bound (primarily albumin), and the remaining 10% complexed with anions, such as sulfate or phosphate. The concentration of calcium distributed among these forms is dependent on the total calcium, the albumin concentration, and the plasma pH. The biologically active form, the free ionized fraction, is tightly regulated by the calcium-regulating hormones: PTH and 1,25-dihydroxyvitamin D (1). PTH directly increases calcium reabsorption in the distal nephron of the kidney through TRPV5 (transient receptor potential vanilloid 5) and also increases the expression of various calcium transport proteins. In response to increased PTH, 1α-hydroxylase in the kidney begins to convert the major reservoir of vitamin D, 25-hydroxyvitamin D, to the biologically active form, 1,25-dihydroxyvitamin D. The 1,25-dihydroxyvitamin D then stimulates calcium absorption in the small intestines through the VDR-RXR (vitamin D receptor-retinoic acid x-receptor complex) to enhance the expression of the epithelial calcium channels. Further, it is also recognized by its receptor in osteoblasts, increasing the expression of the RANKL (receptor activator of nuclear factor-kB ligand), which then induces proosteoclasts to develop to mature osteoclasts and induce bone resorption. Once adequate circulating free calcium is achieved, negative feedback to the parathyroid glands then reduces PTH synthesis. Calcitonin, secreted by the parafollicular cells of the thyroid gland, also acts on calcium concentrations to balance, and oppose, the effects of PTH.
QUESTIONS TO CONSIDER
What is the differential diagnosis for increased calcium concentrations and what other laboratory tests should be considered?
What are the clinical symptoms of hypercalcemia?
What are the common preanalytic and analytic confounders to reporting accurate calcium concentrations?
In what conditions might it be therapeutic to intentionally increase a patient's calcium concentration?
The spectrum of clinical manifestations of hypercalcemia is wide, ranging from few, if any, symptoms with mild hypercalcemia, to obtundation and coma, if severe (2). Patients with mild hypercalcemia (11–12 mg/dL) may be asymptomatic or may have nonspecific symptoms, such as constipation and fatigue. Moderately hypercalcemic patients (12–14 mg/dL) may suffer from polydipsia, polyuria, anorexia, dehydration, nausea, muscle weakness, and changes in sensorium. In severe hypercalcemia (>14 mg/dL), these symptoms often progress and may ultimately lead to coma (2).
Hypercalcemia can result from increased calcium entry into circulation (via increased bone resorption or intestinal absorption), or as a result of decreased urinary excretion. Primary hyperparathyroidism and malignancy account for 80%–90% of all hypercalcemic patients, with the prior comprising the majority of the hypercalcemic ambulatory population and the latter accounting for up to 65% of hypercalcemic hospitalized patients (3). Differentiating between these 2 disease states is typically straightforward, as malignancy is most often clinically evident by the time it causes hypercalcemia with suppressed concentrations of PTH; in most cases PTHrP (PTH-related peptide) will be increased. In contrast, PTH concentrations are high in primary hyperparathyroidism. Although these 2 disease states account for the vast majority of hypercalcemic patients, other diseases and conditions can cause hypercalcemia and must be considered in the differential diagnosis (2).
CLINICAL LABORATORY INVESTIGATION
The first step in the clinical laboratory investigation of hypercalcemia is to determine whether the increased calcium is true or erroneous. Preanalytical effects include patient preparation and the method of sample collection. An important and common source of preanalytical error is the prolonged use of a tourniquet or venous occlusion during sampling, which may falsely increase measured calcium concentrations. Similarly, fist clenching, venous stasis, and significant posture changes may lead to erroneous results. The collection tube used and order of collection is also important because certain additives such as citrate, oxalate, and EDTA bind calcium and falsely decrease measured calcium concentrations (4).
The most common methods for calcium measurement include ion-specific electrodes, for measuring free calcium, and spectrophotometry, the primary method used to measure total calcium. Free calcium concentrations are strongly influenced by pH, as described by an inverse relationship. This is because ionized calcium (Ca2+) competes with hydrogen ions (H+) to bind negatively charged proteins and anions. As the pH decreases (becomes more acidic), H+ increases and Ca2+ has more competition for these binding sites, thus, the concentration of circulating free Ca2+ increases. Generally, free calcium changes by approximately 0.2 mmol/L for every 0.1 pH unit change (4). Erroneously LOW free calcium can also occur in vitro from underfilled tubes, or tubes uncapped before analysis, because the loss of carbon dioxide increases sample pH (less acidic, increases protein-bound Ca).
There are a number of substances that have been reported to interfere with spectrophotometric methods for the measurement of total calcium including hemolysis, icterus, lipemia, paraproteins, magnesium, and gadolinium in contrast agents (4). While not as accurate as a measured free calcium, total serum calcium concentrations should be corrected for hyper- or hypoalbuminemia. Several equations, some of which are method specific, have been proposed, but are all generally of the following form (4):
Corrected Ca = Total Ca + 0.8(4.0 − Albumin).
RESOLUTION OF CASE
All of the above sources of error were investigated in this case without any identifiable issues. Free calcium results were determined in samples collected and maintained anaerobically until the time of measurement, minimizing pH changes. There was no evidence of spectrophotometric interference since this patient's hemolysis, lipemia, and icterius indicies were below their reporting limits. Instrument results were checked against those resulted and postanalytic sources of error, including reporting errors, did not occur.
A review of the patient's laboratory data revealed clinical concordance with the increased calcium concentrations (Fig. 1). Both the PTH and the phosphate concentrations were correspondingly decreased, illustrating that this patient had intact feedback mechanisms regulating calcium homeostasis and ruling out primary hyperparathyroidism. The calcium concentrations were so high that the possibility of intravenous contamination of the sample was investigated, at which point the laboratory discovered that the patient was receiving an intravenous bolus of calcium chloride solution with a therapeutic goal of significant hypercalcemia.
Further investigation revealed that the patient had called 911 after an accidental ingestion of roughly ten 120-mg pills of her prescribed verapamil, a calcium channel blocker (CCB). She had been transferred from an outside hospital where she presented in cardiogenic shock. On arrival, the patient had a Glasgow Coma Score of 3 and was pulseless despite maximal doses of pressors. She was emergently taken to the catheterization laboratory for a temporary pacemaker implantation and intraaortic balloon pump placement. Her blood glucose was increased at 376 mg/dL and blood gas analysis showed a severe metabolic acidosis (pH: 6.91, pCO2: 28 mmHg, HCO3: 6 mEq/L), and increased lactate (10.7 mmol/L).
Once stabilized, the patient was treated for verapamil overdose and severe metabolic acidosis according to our institutional guidelines for calcium channel blocker overdose. The patient received bolus intravenous doses of calcium chloride to overcome the calcium channel blockade, with a therapeutic goal of 2.0 mmol/L for free calcium. To prevent spurious elevations in the measured calcium due to infusion contamination and/or incomplete electrolyte equilibrium, samples were collected 30 min after administration of the calcium chloride bolus. The patient also received insulin to treat her hyperglycemia, which resulted from the CCB overdose. She eventually stabilized, was successfully extubated, and discharged home in good condition.
CCB OVERDOSE AND THERAPEUTIC HYPERCALCEMIA
Verapamil is a CCB used as an antiarrhythmetic agent and it is particularly useful in the management of supraventricular tachyarrhythmias (5, 6). CCB toxicity is characterized primarily by its cardiovascular effects, which include bradycardia, conduction abnormalities, hypotension, and if severe, cardiogenic shock (7). Patients with CCB overdose may require aggressive initial resuscitation, with many patients presenting with CNS depression and a loss of airway protection reflexes; ultimately requiring intubation and mechanical ventilation. Impaired cardiac contractility and peripheral vascular resistance may necessitate pharmacotherapy with cardiotonic and vasoactive drugs.
In response to stress, increased glycogenolysis increases blood glucose concentrations; however, due to CCB blockade of calcium-mediated insulin release by pancreatic β cells, the additional glucose cannot be used. As with this case, patients can present with clinical manifestations similar to diabetic ketoacidosis, including hyperglycemia, insulin deficiency, and lactic acidiosis (7). These effects may be managed through insulin administration, appropriate ventilator settings, and bicarbonate administration.
In addition to initial resuscitation and supportive care, calcium infusion is the primary recommended treatment for CCB toxicity. Increasing extracellular calcium concentrations is aimed at overcoming the drug's effects on the calcium channel through competitive antagonism and generally increasing calcium entry through the unblocked channels (8). It is recommended to follow the serum free calcium concentration, not only because of faster analysis time in whole blood compared to serum analysis, but also because there are no corrections required for albumin concentration. Interestingly, because this patient's albumin was low, the corrected calcium (18.6 mg/dL) was even higher than the measured total calcium (16.9 mg/dL) initially flagged as a critical result. At that time, her free calcium was 2.2 mmol/L, which, while more than twice the upper limit of the reference interval, was accurate even in the setting of hypoalbuminemia.
In summary, this is a case of severe hypercalcemia in a patient who went into cardiogenic shock after accidental verapamil overdose. After initial resuscitation, the patient was treated with an intravenous infusion of calcium chloride in an effort to overcome the drug's toxic effects by competitive antagonism. The significantly increased free and total calcium concentrations reported from the laboratory were found to be accurate and interestingly therapeutic in nature, rather than reflecting a primary disease state or laboratory error.
POINTS TO REMEMBER
Primary hyperthyroidism and malignancy account for the vast majority of cases of hypercalcemia and can be assessed by considering PTH concentrations.
There are a number of preanalytical errors that can affect the accuracy of free calcium, namely changes in sample pH as a result of underfilled collection tubes, or tubes uncapped before analysis.
CCB toxicity can result in cardiogenic shock and diabetic ketoacidosis-like hyperglycemia due to its effects on the cardiac L-type voltage-gated calcium channels and calcium-mediated insulin release by pancreatic β cells, respectively.
The primary recommended treatment for CCB toxicity includes increasing extracellular calcium to overcome the drug's effects on the calcium channel through competitive antagonism.
In an acute setting of CCB overdose, serum free calcium concentration should be used to monitor therapeutic calcium infusion due to a rapid turnaround time without the need to correct for the serum albumin concentration.
↵2 Nonstandard abbreviations:
- parathyroid hormone;
- ionized calcium;
- hydrogen ions;
- calcium channel blocker.
Author Contributions: All authors confirmed they have contributed to the intellectual content of this paper and have met the following 3 requirements: (a) significant contributions to the conception and design, acquisition of data, or analysis and interpretation of data; (b) drafting or revising the article for intellectual content; and (c) final approval of the published article.
Authors' Disclosures or Potential Conflicts of Interest: Upon manuscript submission, all authors completed the author disclosure form. Disclosures and/or potential conflicts of interest:
Employment or Leadership: G.L. Horowitz, Clinical Chemistry, AACC.
Consultant or Advisory Role: None declared.
Stock Ownership: None declared.
Honoraria: None declared.
Research Funding: None declared.
Expert Testimony: None declared.
Patents: None declared.
- Received for publication May 24, 2016.
- Accepted for publication August 9, 2016.
- © 2016 American Association for Clinical Chemistry