Acta Neuropathologica

, Volume 110, Issue 1, pp 77–83

Cerebellopontine calcification: a new entity of idiopathic intracranial calcification?

Authors

    • Department of Pediatrics and Pediatric NeurologyYokohama Ryo-iku Medical Center
    • Department of PediatricsTokyo Women’s Medical University
  • Makoto Shibuya
    • Department of Laboratory MedicineTokai University
  • Masaharu Hayashi
    • Department of Clinical NeuropathologyTokyo Metropolitan Institute for Neuroscience
  • Shizuko Matsuoka
    • Department of NeurologyYokohama Stroke and Brain Center
  • Kaori Kaneko
    • Department of Pediatrics and Pediatric NeurologyYokohama Ryo-iku Medical Center
    • Department of PediatricsTokyo Women’s Medical University
  • Yuri Chikumaru
    • Department of Pediatrics and Pediatric NeurologyYokohama Ryo-iku Medical Center
  • Kazuyo Saito
    • Department of Pediatrics and Pediatric NeurologyYokohama Ryo-iku Medical Center
  • Akira Matsui
    • Department of Pediatrics and Pediatric NeurologyYokohama Ryo-iku Medical Center
  • Seiji Kimura
    • Department of Pediatrics and Pediatric NeurologyYokohama Ryo-iku Medical Center
Case Report

DOI: 10.1007/s00401-005-1011-y

Cite this article as:
Saito, Y., Shibuya, M., Hayashi, M. et al. Acta Neuropathol (2005) 110: 77. doi:10.1007/s00401-005-1011-y

Abstract

We report the autopsy case of a 40-year-old woman with severe intellectual and motor disabilities, who showed calcification in the cerebellum and pons but not in the basal ganglia on CT scan, and died of intracranial hemorrhage due to intractable hypertension. At autopsy, numerous calcium deposits were noted in the cerebellar cortex, the dentate nucleus, the cerebellar white matter and the ventral pons. These deposits were distributed both in the neuropil and the white matter, but rarely within the arterial walls or in contact with capillaries. This weak relationship between calcification and the blood vessels, in addition to the paucity of basal ganglia calcification, is in contrast to the findings with other disorders involving intracranial calcification, including Fahr’s disease and calcium metabolism disorders. Immunohistochemistry revealed intense staining of calbindin-D28K and parvalbumin at sites of calcium deposits both in the present case and in a case of pseudohypoparathyroidism, whereas these proteins were not localized to calcium deposits in the cerebellum of a Fahr’s disease brain. We propose that the present case may represent a distinct entity among diseases characterized by idiopathic intracranial calcification. In addition, calcium-binding proteins may be involved in the calcification process in some cases with intracranial calcification.

Keywords

CalcificationCalcium-binding proteinCerebellumImmunohistochemistryPons

Introduction

Intracranial calcification, frequently observed in the basal ganglia, accompanies many neurological diseases [4, 11, 14, 15, 18, 25, 35, 37, 42, 44] and calcium metabolism disorders including hypoparathyroidism [10, 12, 29] and pseudohypoparathyroidism [16]. Fahr’s disease is one term used to describe idiopathic cases involving intracranial calcification without any primary disease [26, 27, 39]. In such conditions, basal ganglia calcification (BGC) is pathologically characterized by calcium deposits within or adjacent to the vessel walls, which show no arteriosclerotic changes [14, 28, 38]. Certain diseases with BGC show calcification in other sites of the central nervous system. These include the subcortical white matter [21, 26], the cerebellar cortex and the dentate nucleus [16, 29, 38], and the pons [8, 30]. Microscopically, the calcification in these sites appears the same as seen with BGC. In this report, we describe a patient showing calcification in the cerebellum and pontine nucleus, similarly to some cases of Fahr’s disease [30] and pseudohypoparathyroidism [16]. However, our patient did not show BGC on a computed tomography (CT) scan, and the calcium deposits appeared to have a rare relationship to the blood vessels, in contrast to the aforementioned disorders with BGC. Calcium-binding proteins (CaBPs) are expressed in some subsets of neurons, and modulate their excitability through buffering the intracellular calcium [3]. A distinct distribution of CaBPs in the cerebellum [5] and the brainstem [6], together with the role of CaBPs in the mineralization process of extracerebral organs [9, 17], prompted us to examine the expression of CaBPs at calcification sites of the present case, as well as in Fahr’s disease and pseudohypoparathyroidism brains. We therefore examined the involvement of CaBPs in calcification in the present case and compared it to other disorders with BGC.

Case report

The patient was born to unrelated parents at 38 gestational weeks, after a pregnancy complicated by toxemia at 6 months of pregnancy. Her elder sister’s son has intellectual disabilities, but further information was not available. Her birth weight was 1,600 g and there was no perinatal asphyxia. She showed severe psychomotor developmental delay during infancy, and did not gain the ability to control her head until 1.5 years, and to stay sitting until 3.5 years of age. Epileptic seizures appeared at 15 years of age, and she lost the ability to maintain a sitting position around this period. At 19 years of age, retinitis pigmentosa and atrophy of the retina and choroidea were diagnosed, and a brain CT scan detected no intracranial calcification. Screening examination of a urinary sample for metabolic disorders was negative. She did not experience any further episodes of anoxia or prolonged epileptic seizures until she was admitted to our institute at 26 years of age. Brain CT revealed calcification in the deep cerebellar hemispheres, the cerebellar vermis, and the center of the pontine base, but not in the basal ganglia (Fig. 1A, B). Epileptic seizures were controlled well by administration of carbamazepine, and her health remained good during her thirties. Upon physical examination, she was bed-ridden, and her upper extremities showed some purposeful movements including grasping of toys and tapping on her chest. Her lower extremities showed contracture in the frog-leg posture. Muscle tonus was mildly elevated, deep tendon reflex was normal, and pyramidal tract sign was negative. Blinking to finger thrust, light reflex and acoustic blink were positive at 37 years of age, but had disappeared at 40 years of age. When she was 39, hypertension of 150–180/100–120 mmHg gradually appeared. Plasma renin activity, serum aldosterone, urinary excretion of catecholamines were all in normal range. An analysis of calcium metabolism, including serum parathyroid hormone, calcitonin, osteocalcin, 1,25-dihydroxy vitamin D, and urinary calcium excretion, showed no abnormalities. Serum lactate and pyruvate were normal. The hypertension remained uncontrolled despite administration of depressor drugs, and she died of a massive left thalamic hemorrhage at 40 years of age.
Fig. 1

A, B Brain CT of the present case. A Calcification is observed in the cerebellar vermis, hemispheres and in the pons. B There is no calcification in the basal ganglia. C Gross photographs of the patient’s brain. Left thalamic hemorrhage penetrates into the left lateral ventricle. DG Calcification in the cerebral white matter (D, E) and the basal ganglia (F, G) in the present case (D, F), and the cases of Fahr’s disease (E) and pseudohypoparathyroidism (G). DG Hematoxylin and eosin staining, ×200

Materials and methods

After macroscopic inspection, tissues obtained at autopsy were fixed in 10% buffered formalin, and several regions of the cerebrum, cerebellum, midbrain, pons and medulla oblongata were excised, and embedded in paraffin. Brains from a 54-year-old woman with Fahr’s disease and a 46-year-old woman with pseudohypoparathyroidism were examined as disease controls with the consent of their families. Specimens were sectioned at 4 μm and the sections were stained with hematoxylin and eosin, Klüver-Barrera and von Kossa staining. To characterize the nature and distribution of calcium deposits, immunohistochemical analyses were performed on serial sections of the cerebral cortex, lenticulate nucleus, and the cerebellar cortex with the dentate nucleus and pons, using mouse monoclonal antibodies against glial fibrillary acidic protein (GFAP) (Dako Cytomation, Carpinteria, CA), CD34 (Nichirei, Tokyo, Japan), calbindin-D28K (Sigma-Aldrich, St. Louis, MO), and parvalbumin (Sigma-Aldrich). The sections were deparaffinized, immersed in 1% hydrogen peroxide, and rinsed with TRIS-buffered saline (pH 7.6). Microwave treatment was performed to retrieve each antigen. The sections were incubated with the primary antibodies (GFAP 1:100, CD34 prediluted by the manufacturer, calbindin-D28 K 1:100, parvalbumin 1:100) for 48 hours at 4 °C. Antibody binding was visualized by the avidin-biotin-peroxidase method following the manufacturer’s protocol (Nichirei).

Results

General autopsy findings

The lungs demonstrated multiple bullae. Occasional glomerular sclerosis was present in the kidney. Parathyroid glands were normal in size and histological appearance. A calculus of 7×5×5 cm was present in the vagina. Hypertension-related thickening of the arterial media was found in the heart, kidney, pancreas, and brain. Arteriosclerotic change was not evident in any organs.

Gross pathological findings of the brain

The brain was small and weighed 710 g before fixation. The cerebral cortex was atrophied with occipital predominance. Hemorrhage from the left thalamus penetrated into the lateral ventricle and occupied the whole ventricular system (Fig. 1C). Another focal hemorrhage was serially observed from the ventral midbrain to the rostral pontine tegmentum. Otherwise no abnormality was noted in the cerebellum or the brainstem. Calcification was not discernible on macroscopic examination.

Microscopic findings of the brain

Since massive hemorrhage devastated the left cerebral hemisphere, only the right cerebral hemisphere was subjected to reliable histological observation. In the cerebral cortex, marked loss of neurons with gliosis was noted in the occipital lobe. Myelin loss and a decrease in volume were noted in the periventricular white matter at the posterior horn of the lateral ventricle. Several small calcium concretions were observed in the periventricular white matter (Fig. 1D) and in the deep layers of the temporal cortex, but these had no relationship to the vessels, in contrast to the aggregation of mineral deposits along the capillaries in the same regions of the Fahr’s disease (Fig. 1E) and pseudohypoparathyroidism brains. In the basal ganglia, neuronal loss and gliosis were prominent in the globus pallidus. A few small calcium deposits were present in the thickened arterial media (Fig. 1F), whereas vascular calcification was prominent and accompanied with parenchymal spherites in the disease controls (Fig. 1G). A lacunae infarct was observed in the lateral putamen with infiltrating macrophages.

Neuronal loss in the cerebellar cortex was observed predominantly at the periphery of the hemispheres. Numerous calcium deposits were present in the folia and adjacent white matter of the deep hemispheres (Fig. 2A), and in the dentate nucleus. These concretions were distributed predominantly in the molecular layer, the Purkinje cell layer and the superficial granular layer, again with little relationship to the cortical vessels (Fig. 2C), in contrast to those in the Fahr’s disease (Fig. 2D) and the pseudohypoparathyroidism brains. The same tendency was also observed in the neuropil of the dentate nucleus (Fig. 2E). Granular deposits were found occasionally in the vessel wall or perivascular area (Fig. 2F). These deposits were solitary and distinct in contour, while they were fine, hazy and fusing with the vessel walls in the disease controls (see Fig. 1G). The proportion of calcium deposit-positive vessels was extremely low in this case compared to the disease controls, whereas the total degree of calcification in the cerebellar cortex and the dentate nuclei was comparable. In the pons, small calcium deposits were scattered at the center of the pontine base, involving both the gray (Fig. 2B) and white (not shown) matter. Calcification of the blood vessel walls was absent in this area. A mild loss of neurons and myelinated fibers was found in the inferior olivary nucleus and the posterior funiculus of spinal cord, respectively.
Fig. 2

Calcium deposits in the cerebellar cortex (A, C, D), pontine nucleus (B), dentate nucleus (E) and cerebellar white matter (F) in the present case (AC, E, F) and the case of Fahr’s disease (D). A, B Klüver-Barrera staining, C von Kossa staining, DF hematoxylin and eosin staining; A ×10, B ×50, C ×100, DF ×200

Immunohistochemistry

Immunohistochemistry for CD34, a marker of the endothelium, further characterized the rarity of contact between the calcium deposits and blood vessels in the present case (Fig. 3A), in contrast to the findings in the disease controls (Fig. 3B). The antibody staining for GFAP visualized reactive gliosis surrounding each concretion to a similar extent in all the cases examined (not shown). The Purkinje cells, a proportion of fibers in the cerebellar white matter and the neurons in the dentate nucleus were all immunoreactive for both calbindin-D28K and parvalbumin in all three disease cases. Parvalbumin was also localized by immunostaining in the pontine nucleus neurons. Furthermore, the calcium deposits were immunopositive for both calbindin-D28K and parvalbumin in the cerebellar cortex, the dentate nucleus and the pons in the present case (Fig. 3C, E). Similarly, the deposits were immunopositive for CaBPs in the basal ganglia in both the disease controls (not shown), in addition to the cerebellum in the case of pseudohypoparathyroidism. In contrast, the calcium deposits were not immunoreactive for CaBPs in the cerebellum from the Fahr’s disease brain (Fig. 3D, F). The intensity of immunolabeling for CaBPs varied depending on the brain region in each case, as summarized in Table 1.
Fig. 3

Immunohistochemistry in the cerebellum of the present case (A, C, E) and the case of Fahr’s disease (B, D, F). A, B CD34 is localized to the capillary endothelium. Calcium deposits, counterstained with hematoxylin, are located in the vicinity of the capillaries in the case of Fahr’s disease (B), but are not in contact with vessels in the present case. Arrows in B indicate small capillary walls, not discernible on hematoxylin and eosin staining, at the center of or adjacent to calcium deposits. Inset in B shows calcium deposits surrounding capillaries, at a higher magnification. C, D Calbindin-D28K immunostaining for calcified deposits. Calcium deposits are intensely immunostained in the cerebellar cortex (C), the dentate nucleus and the pons in the present case. In the case of Fahr’s disease, calcium deposits lack immunoreactivity in the cerebellum (D: calcified vessel in the white matter). E, F Parvalbumin immunostaining for calcified deposits. Calcium deposits are immunostained for parvalbumin in the cerebellar cortex (E) in the present case. In the case of Fahr’s disease, calcium deposits (F) are immunonegative in the cerebellum. A, B ×100; CF ×200

Table 1

Immunoreactivities in calcium deposits for calcium-binding proteins ( absent, ± equivocal, + weak, ++ moderate, +++ intense, DN dentate nucleus, N lacking calcium deposits, WM white matter)

Calbindin-D28K

Cerebellum

Pons

Basal ganglia

Parvalbumin

Cerebellum

Pons

Basal ganglia

Cortex

WM

DN

Cortex

WM

DN

Present case

+++

++

+++

++

N

+∼+++

+

+++

+++

N

Fahr’s disease

−∼±

N

++∼+++

−∼±

−∼±

N

+∼+++

Pseudo-

hypoparathyroidism

++

±∼++

+++

N

+∼+++

±∼++

−∼++

±∼++

N

++∼+++

Discussion

Some brain lesions in the reported case may have resulted from an intrauterine hypoxia, and from the accompanying hypertension. These include the neuronal loss and/or gliosis in the cerebral cortex, the periventricular white matter, and the basal ganglia. Such pathological changes may have caused the profound intellectual and motor disabilities, as well as epileptic seizures of the present patient. The small number of calcium deposits in these areas may be secondary to the destructive insults. However, the calcifications in the cerebellum and the pons could not be explained by the above complications, because: (1) the calcification emerged on CT scan during adulthood, (2) neurons and other structures adjacent to the deposits were not damaged in these areas, and (3) the neuronal loss in the cerebellar cortex, possibly secondary to an hypoxic event, was marked at the periphery of the cerebellar hemispheres, whereas the calcification was prominent at the folia located deep within the hemispheres.

Among conditions with cerebellar calcifications (Table 2), pontine calcification is observed only in rare cases of Fahr’s disease [30], calcium metabolism disorders [16], and Aicardi-Goutieres syndrome [24]. However, all of these cases are accompanied by BGC, which was not evident in the present case, with the exception of a few vascular deposits. In diseases with BGC, calcium deposits are usually distributed in the media and adventitia of medium-sized or small vessels, as small calcospherites along the capillaries, and freely in the parenchyma [11, 25, 28, 38]. Unknown or inflammatory processes may result in dysfunction of the vascular endothelium with increased permeability [28], leading to the development of concretions, which could be a mixture of polysaccharides, glycoproteins, mucopolysaccharides, calcium salts, iron and other minerals [22, 26, 38]. Contrary to this hypothesis, the calcium deposits had little relation to the vasculature in the present case.
Table 2

Disorders showing calcification in the cerebellum and/or pons

Cerebellum

Idiopathic [1, 23]

Fahr’s disease [21, 22, 26, 38]

Hypoparathyroidism [12, 16, 29]

Pseudohypoparathyroidism [16]

Aicardi-Goutieres syndrome [15, 24]

Cockayne syndrome [15]

Tuberous sclerosis [40]

Congenital anomalies [45]

Krabbe disease [43]

Cerebrotendinous xanthomatosis [43]

Central lupus [35]

Tumors [32, 41]

Hematoma [19]

Congenital/neonatal infection [4, 31]

Syphilis [20]

Lead intoxication [36]

Pons

Fahr’s disease [30]

Hypoparathyroidism/ Pseudohypoparathyroidism [16]

Moebius syndrome [7, 13]

Multifocal necrotizing leukoencephalopathy [2]

Irradiation [33]

Aicardi-Goutieres syndrome [24]

To explore the pathogenic mechanism of calcification in the present case, we performed immunohistochemistry to examine the distribution and expression of CaBPs. Interestingly, calbindin-D28K and parvalbumin co-localized with the calcium concretions. We assume that this result was not artifactual, based on the variability of staining intensity amongst brain regions, and the absence of immunostaining in the cerebellum of the Fahr’s disease brain, as well as the lack of immunostaining with other mouse monoclonal antibodies against CD34 and GFAP. CaBPs in the cell bodies and neurites of the cerebellar and pontine neurons might be involved in the calcification process that is not related to the blood vessels. However, the presence of CaBPs in the calcium deposits in the disease-control basal ganglia, as well as in the cerebellum in the case of pseudohypoparathyroidism, suggests that this process is not specific to the present case. Further investigations are necessary to elucidate the involvement of CaBPs during intracranial calcification.

A report by Puvanendran and Wong [34] described juvenile hypertension in siblings with idiopathic BGC. The significance of hypertension in the present case remains unknown, but this complication might imply a common pathophysiology among certain cases with intracranial calcifications.

In conclusion, we reported a patient with pontocerebellar calcification. The calcium deposits were characteristic in their anatomical distribution and in the paucity of relationship to the blood vessels. Although the pathomechanism of calcification remains unclear, this type of calcification may represent a distinct entity among diseases involving intracranial calcification.

Copyright information

© Springer-Verlag 2005