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  • Histopathological alterations of the parathyroid glands in haemodialysis patients with secondary hyperparathyroidism refractory to cinacalcet hydrochloride

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Original article
Histopathological alterations of the parathyroid glands in haemodialysis patients with secondary hyperparathyroidism refractory to cinacalcet hydrochloride
  1. Keiichi Sumida1,
  2. Michio Nakamura1,
  3. Yoshifumi Ubara1,
  4. Yuji Marui1,
  5. Kiho Tanaka1,
  6. Kenmei Takaichi1,
  7. Shinji Tomikawa1,
  8. Naoko Inoshita2,
  9. Kenichi Ohashi2
  1. 1Nephrology Center, Toranomon Hospital, Tokyo, Japan
  2. 2Department of Pathology, Toranomon Hospital, Tokyo, Japan
  1. Correspondence to Dr Michio Nakamura, Nephrology Center, Toranomon Hospital, 2-2-2 Toranomon Minato-ku, Tokyo, Japan; mnakamura{at}toranomon.gr.jp

Abstract

Background Cinacalcet treatment for secondary hyperparathyroidism (SHPT) has demonstrated parathyroid size regression and morphological changes, such as cystic degeneration and hypovascularisation, on ultrasonography.; However, there have been very few reports regarding the histopathological alterations of hyperplastic parathyroid glands in patients with SHPT after administration of cinacalcet. The aim of this study was to elucidate the effects of cinacalcet for histopathological alterations on the parathyroid glands.

Methods A total of 92 hyperplastic parathyroid glands were obtained from 24 dialysis patients with severe SHPT who underwent total parathyroidectomy and were enrolled in this study. Patients were divided into those treated with and without cinacalcet (cinacalcet group and conventional group, respectively; both n=12). The areas of oxyphil cells, cystic degeneration, haemorrhagic changes and haemosiderin deposition were assessed semiquantitatively.

Results Total maximal parathyroid gland weight and maximal-to-minimal parathyroid gland weight ratio were significantly higher in the cinacalcet group compared with the conventional group (1798.7±1658.3 mg vs 764.2±471.1 mg, p=0.018, 15.8±13.9 vs p=0.047, 6.6±4.2, respectively). Significant increases were observed in oxyphil cell area (61.7%±17.1% vs 36.7%±15.6%, p=0.001) and haemosiderosis score (1.50±1.24 vs 0.42±0.51, p=0.029) in the former rather than the latter group.

Conclusions These results suggest that cinacalcet may induce specific qualitative alterations of hyperplastic parathyroid glands in patients with severe SHPT.

  • Secondary hyperparathyroidism
  • cinacalcet hydrochloride
  • parathyroid gland
  • histopathological alteration
  • nephrology
  • parathyroid
  • histopathology
  • colorectal cancer
  • gall bladder
  • oncogenes
  • p53
  • pancreas
  • immunohistochemistry
  • renal

https://doi.org/10.1136/jclinpath-2011-200100

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    Introduction

    Secondary hyperparathyroidism (SHPT) is one of the most important complications in chronic kidney disease—mineral and bone disorders. Patients are considered to have severe SHPT when serum phosphorus (P), calcium (Ca), the calcium–phosphorus product (Ca–P product) and intact parathyroid hormone (PTH) levels can no longer be adequately controlled by medical management and when clinical symptoms are associated with a significantly increased risk of cardiovascular morbidity and mortality.1–4 Despite the recent development of novel therapeutic modalities, including new phosphate binders and new vitamin D analogues, severe SHPT remains uncontrollable in some patients. The Japanese Society of Dialysis Therapy guidelines proposed that parathyroid intervention therapy, including parathyroidectomy (PTx), should be performed in such patients with persistent high serum levels of intact PTH >500 pg/ml, associated with hyperphosphataemia and/or hypercalcaemia that are refractory to medical management.5

    Cinacalcet hydrochloride (cinacalcet), a calcimimetic, is a new agent for SHPT treatment, which increases the sensitivity of calcium-sensing receptors to activation by extracellular calcium and thus suppresses PTH release, while simultaneously controlling parameters associated with Ca–P balance.6–9 Cinacalcet has the potential to control biochemical parameters even in cases of severe SHPT refractory to conventional treatment with calcium supplements, phosphate binders and active vitamin D analogues.

    A recent study demonstrated parathyroid size regression and morphological changes, such as cystic degeneration and hypovascularisation, during cinacalcet treatment.10 However, these findings were observed only after ultrasonography, and there have been very few reports to date regarding the histopathological alterations of hyperplastic parathyroid glands in patients with SHPT after administration of cinacalcet.11

    To elucidate the effects of cinacalcet on the parathyroid glands, we performed histopathological analyses of the hyperplastic parathyroid glands removed from haemodialysis patients with severe cinacalcet–refractory SHPT, together with examination of biochemical parameters.

    Methods

    Patients

    A total of 24 dialysis patients with severe SHPT who underwent total PTx with autotransplantation into the forearm muscle at our institute from January 2008 to February 2010 were enrolled in this study. The indications for PTx of SHPT were based on clinical and/or laboratory data according to the guidelines of the Japanese Society for Dialysis Therapy.5

    The patients were divided into two groups according to the medical treatment for SHPT with or without cinacalcet. The cinacalcet group consisted of 12 patients (44 hyperplastic parathyroid glands) treated with cinacalcet in combination with conventional medical treatment, such as calcium supplements, phosphate binders and active vitamin D analogues, while the conventional group consisted of 12 patients (48 hyperplastic parathyroid glands) given only conventional treatment. The mean duration of cinacalcet treatment was 16.1±6.8 months, and the mean dose of cinacalcet was 64.6±39.1 mg/day.

    Biochemical analysis

    Blood samples of these patients were obtained within 1 week prior to PTx. Serum albumin, calcium, phosphorus, alkaline phosphatase, osteocalcin and intact PTH were measured using standard methods. The measured serum calcium levels were adjusted to albumin levels using the following equation: corrected calcium=serum measured calcium+(4–albumin). This study was performed in accordance with the principles of the Declaration of Helsinki and was approved by the ethical committee of Toranomon Hospital. All patients provided informed consent prior to enrolment in the study.

    Histological studies of parathyroid gland tissues

    A total of 92 hyperplastic parathyroid glands were obtained from the 24 haemodialysis patients enrolled in this study. Four normal parathyroid glands resected together with four benign thyroid tumours (two adenomatous goitres and two follicular adenomas) were obtained from four patients with normal renal function and were used as positive controls. Each parathyroid gland removed was weighed just after PTx. Histopathological studies of the parathyroid glands with light microscopy were performed by the same pathologist in a blinded manner. We evaluated the largest hyperplastic parathyroid gland obtained from each patient in this study. Hyperplasia was classified as either diffuse or nodular hyperplasia. Diffuse hyperplasia was defined as an increased number of parenchymal cells with normal lobular structures, and nodular hyperplasia was defined as at least one well-circumscribed, encapsulated and virtually fat cell-free accumulation of parenchymal cells.12

    Haemorrhagic changes were defined as haemorrhage filled with red blood cells observed inside the parenchyma of hyperplastic nodules. Haemosiderin deposition was mostly observed in macrophages, especially around haemorrhagic changes in hyperplastic nodules. These findings were assessed semiquantitatively as described below.

    Semiquantitative analysis

    Semiquantitative analyses were performed to assess the histopathological changes. On histopathological analysis, the percentage of oxyphil cell area to total area was evaluated in each of the largest parathyroid gland at a magnification of ×200. Cystic degeneration, haemorrhagic changes and haemosiderin deposition observed in the largest parathyroid gland were also assessed semiquantitatively. Each area, counted at a magnification of ×40 or ×100, was scored according to the percentage of total area using the following grading criteria: grade 3, >50%; grade 2, 25%–50%; grade 1, <25%; and grade 0, no lesion (figure 1). These were designated as the cystic score, haemorrhagic score and haemosiderosis score, respectively. These analyses were performed by three independent observers.

    Figure 1
    Figure 1

    Histopathological changes of hyperplastic parathyroid gland. (A–C) Cystic degeneration (original magnification ×40). (D–F) Haemorrhagic change (original magnification ×40). (G–I) Haemosiderin deposition (original magnification ×100). Each area was scored according to the percentage of total area using the following grading criteria: grade 3, >50% (A, D, G); grade 2, 25%–50% (B, E, H); grade 1, <25% (C, F, I); and grade 0, no lesion.

    Statistical analysis

    All values are expressed mean±SEM. Statistical significance was determined using the χ2 test for categorical variables, Student t test for parametric variables and Mann–Whitney U test for non-parametric variables for comparison between the two groups. In all analyses, which were performed using the SPSS software package (SPSS 11.01J for Windows; SPSS Inc.), p<0.05 was considered statistically significant.

    Results

    Patient characteristics

    Table 1 shows the demographic, clinical and biochemical characteristics of the entire group of 24 patients. Table 2 shows the comparison of clinical, biochemical and histopathological data between the cinacalcet group (n=12) and the conventional group (n=12). There were no significant differences in age, gender or duration of haemodialysis between the two groups. In the cinacalcet group, the mean administered dose and duration of treatment with cinacalcet were 64.6±39.1 mg/day and 16.1±6.8 months, respectively. The mean administered dose of calcitriol in the cinacalcet group was lower than that in conventional group (1.2±1.2 vs 2.5±2.2 μg/wk, p=0.130). In the cinacalcet group, preoperative serum-corrected Ca level was significantly lower than that in the conventional group (9.8±0.7 vs 10.6±0.9 mg/dl, p=0.036).

    View this table:
    Table 1

    Demographic, clinical and biochemical characteristics of the 24 patients

    View this table:
    Table 2

    Comparison of clinical, biochemical and histopathological data between the two groups

    Both the weight of the maximal parathyroid gland and the weight ratio of maximal-to-minimal parathyroid glands were significantly higher in the cinacalcet group than in the conventional group (1798.7±1658.3 mg vs 764.2±471.1 mg, p=0.018; 15.8±13.9 vs 6.6±4.2, p=0.047, respectively). Although the differences between the two groups were not significant, preoperative levels of serum intact PTH tended to be lower, and both the total parathyroid gland weight and the maximal-to-total parathyroid gland weight ratio tended to be higher in the cinacalcet group in comparison with the conventional group.

    Histopathological characteristics of removed parathyroid glands

    A total of 92 hyperplastic parathyroid glands from 24 dialysis patients with SHPT and four normal parathyroid glands from four patients with normal kidney function were obtained in the present study. All normal parathyroid glands showed clusters of parenchymal cells admixed with a considerable amount of adipose tissue (figure 2A). In the cinacalcet group, the specimen of the largest hyperplastic parathyroid gland showed interstitial atrophy with oedema around well-circumscribed and encapsulated chief cell areas and many hyperplastic nodules of oxyphil cells (figure 2B).

    Figure 2
    Figure 2

    Histopathological features of normal and hyperplastic parathyroid glands in the cinacalcet group (H&E staining, original magnification ×40). (A) Normal parathyroid gland showing clusters of parenchymal cells admixed with a considerable amount of adipose tissue. (B) Nodular parathyroid hyperplasia showing interstitial atrophy with oedema around well-circumscribed and encapsulated chief cell areas (arrow) and enlarged nodular hyperplasia composed of oxyphil cells (arrowheads).

    In hyperplastic parathyroid glands, some specific histological changes, such as increased number of oxyphil cells, cystic degeneration, haemorrhagic changes and haemosiderin deposition, were observed by light microscopy, and these changes were markedly increased in the cinacalcet group (figure 1). Cystic degeneration varied in size and was observed only in areas of nodular hyperplasia.

    In semiquantitative analysis, the percentage of oxyphil cell area to the total area was significantly higher in the cinacalcet group compared with the conventional group (61.7%±17.1% vs 36.7%±15.6%, p=0.001) (figure 3A). Haemosiderosis score was increased significantly in the cinacalcet group compared with the conventional group (1.50±1.24 vs 0.42±0.51, p=0.029) (figure 3D), although there were no significant differences between the two groups in either cystic or haemorrhagic score (figure 3B–C).

    Figure 3
    Figure 3

    Comparison of histopathological changes between the two groups. (A) Oxyphil cell area. (B) Cystic score. (C) Haemorrhagic score. (D) Haemosiderosis score. The percentage of oxyphil cell area to the total area was significantly higher in the cinacalcet group compared with the conventional group (61.7%±17.1% vs 36.7%±15.6%, p=0.001) (A). There were no significant differences between the two groups in either cystic or haemorrhagic score (B–C). Haemosiderosis score was increased significantly in the cinacalcet group compared with the conventional group (1.50±1.24 vs 0.42±0.51, p=0.029) (D). The values shown are mean±SEM. *p<0.1; **p<0.05; ***p<0.01.

    Discussion

    Calcimimetics are drugs that mimic or potentiate the action of [Ca2+] at calcium-sensing receptors. Cinacalcet, the first calcimimetic drug approved by the US Food and Drug Administration in 2004, has become available for treatment of dialysis patients affected by SHPT.13–16 Cinacalcet successfully lowers serum intact PTH and simultaneously controls the parameters associated with the Ca–P balance.6 7 Long-term studies have yielded similar results.17 Since its approval for use in Japan in 2008, cinacalcet has been proposed as a new strategy for treatment of SHPT in this country. However, in a minority of patients with cinacalcet treatment, parathyroid overfunction persists, progressively eludes medical control and eventually becomes severe, requiring surgical PTx. There have been few reports regarding the effects of cinacalcet on morphological and pathological variations in hyperplastic parathyroid glands in haemodialysis patients with advanced SHPT.10 11

    First, the clinical features regarding preoperative examinations were characterised. In the patients treated with cinacalcet, the serum-corrected Ca levels were significantly lower and the intact PTH levels were slightly reduced, although not significantly, in comparison with the conventional group. This was considered due to a direct effect of cinacalcet itself. The parathyroid glands from patients receiving cinacalcet treatment showed a number of defining features. Although the values of both intact PTH and serum-corrected Ca were lower in the cinacalcet group, the weight of the maximal parathyroid gland and the maximal-to-minimal parathyroid gland weight ratio were significantly higher compared with those in the conventional group. In general, large parathyroid glands weighing more than 500 mg are characterised histologically by nodular hyperplasia, and a diagnosis of advanced SHPT is made in patients with such glands.18–20 That is, some patients treated with cinacalcet have advanced SHPT histologically, despite low levels of serum Ca and intact PTH. These observations indicate a discrepancy between clinical biochemical parameters and the histopathological severity of this disease, and it is therefore difficult to determine the indications for surgical PTx in some cases. Patients are considered to have advanced SHPT when serum P, Ca, Ca–P product and intact PTH levels can no longer be adequately controlled by conventional therapy, and PTx becomes necessary in such patients in accordance with international guidelines.1 Prolonged progression without surgical intervention represents a risk of enhanced bone resorption relative to formation. Among all the patients administered cinacalcet in the present study, four out of seven patients presenting with normal serum-corrected Ca levels were characterised by high levels of alkaline phosphatase and/or osteocalcin. The reasons for conversion to PTx in the present study included prolonged high levels of bone metabolic parameters (n=8) and increasing size on sonographic images (n=4).

    In turn, we examined whether there was an association between histopathological alterations of parathyroid glands (expression of cell phenotypes and characteristic degeneration) and treatment with or without cinacalcet in patients with refractory SHPT. All of the patients receiving cinacalcet treatment in the present study had more than one nodular hyperplastic gland, considered as the causative nodule for advanced SHPT refractory to cinacalcet. In addition, pathological features in the maximal glands included hyperplasia of oxyphil cells with atrophic areas of chief cells, cystic degeneration, haemorrhagic changes and haemosiderin deposition. Lomonte et al11 reported that oxyphil/chief cell ratio was significantly increasing in the glands of patients treated with cinacalcet and suggested that it was probably due to a more significant decrease in the absolute number of chief cells than because of an increase in the absolute number of oxyphil cells. It was also shown that cinacalcet accelerates chief cell death, probably through an apoptotic mechanism in uraemic rats.21 22 Cystic degeneration of parathyroid glands is found not only in patients treated with cinacalcet but also in those given conventional treatment. However, there was an apparent increase in the area of cystic degeneration. Meola et al10 detected cystic degeneration and hypovascularisation on sonographic images of patients with long-term administration of cinacalcet. In addition, after temporary suspension of cinacalcet treatment, the increase in serum intact PTH values was smaller as compared with the initial PTH levels, possibly because of a reduction in the number of functional parathyroid cells (mass reduction).10 The increase in the area of cystic degeneration found in the present study may represent histopathological corroboration of the phenomena seen on sonographic imaging by Meola et al.10 It is possible that cinacalcet has a qualitative influence on parathyroid cells and induces characteristic pathological changes.

    It should be noted that haemosiderin deposition was significantly increased in the cinacalcet treatment group. Haemosiderin deposition occurs due to interstitial chronic haemorrhage, possibly as a result of microvascular leakage. Meola et al10 also reported diminished vessels of parathyroid glands on sonographic images. Similar phenomena may occur in the microvasculature within the parathyroid glands.

    The main limitations of this study were the small number of patients and that data for the control patients were collected retrospectively. Further studies are required to confirm the findings and gain a better understanding of the underlying mechanisms.

    In conclusion, cinacalcet normalises the levels of biochemical markers of SHPT and is presumed to induce various histopathological alterations in the parathyroid glands. The histological alterations found in the patients treated with cinacalcet include an increased area of oxyphil cells, characteristic cystic degeneration and haemosiderin deposition. However, further studies are required to determine the physiopathological mechanisms underlying glandular degeneration in these patients.

    Take-home message

    Our results suggest that cinacalcet may induce specific qualitative alterations of hyperplastic parathyroid glands in patients with severe SHPT.

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    Footnotes

    • Funding This study was partially funded by the Okinaka Memorial Institute for Medical Research and supported by a grant from The Kidney Foundation, Japan (JFKB 10-8).

    • Competing interests None.

    • Patient consent Obtained.

    • Ethics approval This study was conducted with the approval of the ethical committee of Toranomon Hospital.

    • Provenance and peer review Not commissioned; externally peer reviewed.