Ionized hypercalcemia in 238 cats from a referral hospital population (2009‐2019)

Abstract Background Ionized calcium concentration ([iCa]) is more sensitive for detecting calcium disturbances than serum total calcium concentration but literature on ionized hypercalcemia in cats is limited. Urolithiasis is a possible adverse consequence of hypercalcemia. Hypothesis/Objectives To describe clinical details of diagnoses associated with ionized hypercalcemia in cats and association with urolithiasis. Animals Cats (238) seen between 2009 and 2019 at a referral hospital with [iCa] above the normal reference interval. Methods Observational cross‐sectional study. Signalment, serum biochemical and imaging findings were reviewed for cats with ionized hypercalcemia considered to be clinically relevant (>1.41 mmol/L). Data were summarized by cause of hypercalcemia (i.e., diagnosis). Results Diagnoses for the 238 cats with [iCa] >1.41 mmol/L included: acute kidney injury (AKI; 13%), malignancy‐associated (10.1%), idiopathic hypercalcemia (IHC; 10.1%), chronic kidney disease/renal diet‐associated (8.4%), iatrogenic (5.5%), primary hyperparathyroidism (2.1%), vitamin D toxicity (2.1%) and granulomatous disease (1.7%). In 112 cases (47.1%), no cause for ionized hypercalcemia could be determined (n = 95), hypercalcemia was transient (n = 12), or the cat was juvenile (<1 year; n = 5). Urolithiasis was identified in 83.3% of AKI, 72.7% of iatrogenic, 61.1% of CKD/renal diet‐associated and 50% of IHC cases that were imaged (<50% for other diagnoses). Diagnoses with a high proportion of concurrent total hypercalcemia included primary hyperparathyroidism (100%), vitamin D toxicity (100%), malignancy‐associated (71.4%), granulomatous disease (66.7%) and IHC (65.2%). Conclusions and Clinical Importance Ionized hypercalcemia was most commonly associated with kidney diseases, neoplasia or IHC. The proportion of urolithiasis cases varied by diagnosis.


| INTRODUCTION
Calcium is important for many cellular functions. Although assessment of serum total calcium concentration is available on most routine serum biochemistry panels, ionized calcium (iCa) is the biologically active fraction. 1 Total hypercalcemia has poor sensitivity for detecting ionized hypercalcemia in cats 2,3 and therefore [iCa] should be measured directly whenever possible. Ionized hypercalcemia is caused by a number of underlying pathological and physiological processes, 4 however the relative frequencies of these diagnoses have not been well described. Initial research investigating the causes of hypercalcemia in cats relied on total hypercalcemia. 5 Subsequently, the condition idiopathic hypercalcemia (IHC) was described, 6 which is now often cited as the most common cause of ionized hypercalcemia in cats. 4 A recent study of 119 cats with ionized hypercalcemia did not corroborate this finding, identifying IHC in only 13%. 7 Twenty-three percent of that study population had neoplasia, and a substantial proportion (28.6%) with an [iCa] above their normal reference interval had nonpathologic, transient or inconsequential hypercalcemia. 7 Clinical details of these cats, aside from broad diagnostic groups, are lacking.
Therefore, further data from larger groups of cats are required to better understand the current distribution of diseases leading to ionized hypercalcemia.
Previous studies have associated urolithiasis with both total (11% of hypercalcemic cats) 5 or ionized hypercalcemia (14% of hypercalcemic cats), 8 but the relationship between hypercalcemia and urolithiasis is complex in both humans and animals. 9 Unless associated with urinary tract obstruction and acute kidney injury (AKI), urolithiasis is arguably a consequence rather than a cause of hypercalcemia. Calcium oxalate urolithiasis is associated with hypercalciuria, 10,11 but hypercalcemia is not the sole determinant of hypercalciuria. 9 Urolithiasis previously has been documented only to occur in 35% of cats with IHC 6 and only 14% of cats with obstructive ureteral uroliths are reported to have total hypercalcemia, 12 despite 98% of obstructing ureteroliths being composed of calcium oxalate. 13 Further information on the causes of ionized hypercalcemia and urolith formation in cats therefore is required.
Our aims were to describe cats with ionized hypercalcemia (cases) in a referral center and to describe the biochemical and signalment differences among causes of ionized hypercalcemia (diagnoses). A secondary aim was to identify the proportion of cases with each diagnosis that had concurrent urolithiasis.

| MATERIALS AND METHODS
An observational cross-sectional study was performed. Medical records of all cats presented to the small animal referral hospital of the Royal Veterinary College (RVC) between 1st January 2009 and 1st January 2019 were retrospectively searched using the VetCompass online database to identify all cases of ionized hypercalcemia. Ethical approval had been granted by the RVC Ethics Review Committee (SR20181652).
An iterative process was used to develop a set of effective search terms that could be applied within VetCompass™ for capture of unique candidate cats. The full clinical records then were reviewed manually. Final search terms applied were Hypercal*, Hyperparathyroid*, "ionised calcium", "ionized calcium", iCa, Ca2+, "Ca++", PTH, "parathyroid hormone", "vitamin D", Pamidronate, and "free calcium".
During the study period, whole blood iCa measurements were performed immediately after collection according to manufacturer Each case was reviewed by a resident (SEB) and diplomate (RFG) in internal medicine, and assigned a diagnosis largely based on differential diagnoses previously described for ionized hypercalcemia in cats, 4 using diagnostic criteria presented in Table 1. Diagnoses were retrospectively updated if follow-up data indicated a different disease than that originally assigned, or if no diagnosis had been recorded in the clinical records. A single diagnosis was assigned per cat.
International Renal Interest Society (IRIS) stage and the presence or absence of a SC ureteral bypass (SUB™) system additionally was recorded for patients in the chronic kidney disease (CKD)/renal dietassociated group. 18 The AKI cases were subcategorized by cause (obstructive versus nonobstructive).   Table 2 as "undetermined" and the distribution of [iCa] is represented in Figure 1. They then were removed from further statistical analyses.
Physiological hypercalcemia associated with growth was consid- Clinicopathological data for 138 cats where a diagnosis for their ionized hypercalcemia was reached are presented in Table 4.
Twelve (5%) cases had resolution of the hypercalcemia without specific intervention and were designated as having transient hypercalcemia. Only 1/11 had concurrent total hypercalcemia. The median follow-up for this group was 61 days (IQR, 8.5-508.5). The magnitude of ionized and total calcium concentrations was lower in this group compared to a number of other etiologies (see Table 4). The distribution of first and maximal [iCa] for each diagnosis is depicted in Figure 1.
The correlation between ionized and total calcium concentrations, separated by diagnosis, is depicted in Figure 2. as previously reported (3%-5.6%). 5,7,8 Cats with primary hyperparathyroidism had the highest median age at presentation (15.5 years), similar to a prior case series, 22 suggesting that primary hyperparathyroidism is largely a disease of middle-aged to older cats, as seen in dogs, 23,24 albeit occurring at a much lower frequency.
Only 34 (14.3%) of all cats with ionized hypercalcemia in our study were referred for investigation of hypercalcemia (total or ionized), along with other clinical findings, but 80% of cats with toxicity as a cause of their hypercalcemia had hypercalcemia as a presenting complaint.
Concurrent total hypercalcemia was identified for all diagnoses, and a significant difference in the severity of total hypercalcemia was identified between many of these diagnoses, likely explaining the discrepancy with previous studies that focused on total hypercalcemia. 5 Low proportions of total hypercalcemia were identified in cases with transient ionized hypercalcemia (9.1%), iatrogenic hypercalcemia (30%), AKI (40.7%) and CKD (50%). Nearly 60% of AKI and 50% of CKD cases therefore would not have been included in previous studies. 5 In contrast, malignancy-associated, primary hyperparathyroidism and toxicity had high percentages of cases (>70%) with concurrent total hypercalcemia.
Idiopathic hypercalcemia (IHC) was first described in 2000 in cats from a number of university and private veterinary practices, and the signalment and clinical signs in our study are similar to those previously reported. 6 Only 12.5% of the current IHC cases were presented with polyuria and polydipsia (PU PD). This observation is consistent with previous reports of low frequency of PU PD in hypercalcemic cats, 6 and in contrast to what is reported in dogs (43%-62%). [23][24][25] The proportion of total hypercalcemia in cases of IHC has not previously been reported. We determined that nearly 35% of cats with IHC (where measured) in our study population did not have concurrent total hypercalcemia and therefore IHC cannot be excluded based on the presence of total normocalcemia. A similar proportion of cats was diagnosed with IHC in our study (10.1%) as compared with the study examining ionized hypercalcemia in cats presented to Cornell University Hospital for Animals (13%), 7 which is in contrast to the general perception regarding the frequency of this condition in cats with hypercalcemia (i.e., that IHC is the most common cause of ionized hypercalcemia in cats). 4 It is possible that the cats in our study that had an undetermined cause for their ionized hypercalcemia also could have had IHC and therefore IHC may have been more common than is reported here. Alternatively, some cases may have been misclassified as IHC when another cause for ionized hypercalcemia was present. Regardless, we found IHC to be 1 of the 4 most common diagnoses for cats with ionized hypercalcemia. . The association between AKI and hypercalcemia is complex and, despite strict definitions, some cats may have been allocated to the AKI group when another cause for ionized hypercalcemia also may have been present. It is also possible that breed and neuter status were incorrectly recorded in the hospital computer system. The selection of cases from a referral hospital likely introduced bias, and we may have identified relatively fewer cases of conditions routinely identified and treated in a primary care setting.
We adjusted for the use of 3 different in-house machines for [iCa] measurement by applying one hospital-derived reference range to all results. The upper limit of this calculated reference range was similar to a previously derived in-house reference range for the iSTAT 15 and the external laboratories utilized during the study period.
However, the use of multiple machines remains a limitation of our study. A further limitation is that 2 different biochemistry analyzers were used for total calcium measurement during the study period, but the bias between the machines was minimal and the reference range at the time of sampling was used to define total hypercalcemia, minimizing the impact on our findings.
Another major limitation was the use of the first increased [iCa] from which a diagnosis was made. In conclusion, AKI, CKD, IHC and malignancy-associated ionized hypercalcemia all were found to be causes of clinically relevant ionized hypercalcemia. In our study, the number of cases of transient hypercalcemia and the discrepancy between machine-and hospitalspecific reference intervals for [iCa] highlight the importance of confirming the persistence and clinical relevance of ionized hypercalcemia before embarking on intensive diagnostic testing, ideally using a reference interval derived specifically for the hospital and machine in use.
The proportions of total hypercalcemia for the different diagnoses also emphasize the risk of relying purely on total calcium to determine a cat's calcium status. Uroliths commonly were identified during imaging of hypercalcemic cats in our study and many were not obstructive.
Therefore, abdominal imaging should be considered in all cats with persistent ionized hypercalcemia.