Loss of heterozygosity and somatic mutations of the VHL tumor suppressor gene in sporadic cerebellar hemangioblastomas.

Cerebellar hemangioblastoma is a benign central nervous system neoplasm with characteristic proliferation of vascular and stromal cells. There is increasing evidence that the stromal cell population may represent the neoplastic component of hemangioblastoma, whereas the vascular component may be composed of reactive, nonneoplastic cells. Therefore, successful genetic testing for loss of heterozygosity requires selective analysis of target cell populations. Here, tissue microdissection was used to selectively analyze the stromal cell component of 20 archival sporadic cerebellar hemangioblastomas for loss of heterozygosity at the Von-Hippel Lindau (VHL) gene and somatic VHL gene mutations. Allelic deletions at the VHL gene locus were detected in the stromal cell component with one or more markers (D3S1038, D3S1110, and/or 104/105) in 10 of 19 (52.6%) informative cases. In all cases, heterozygosity at the VHL gene locus was retained in the vascular component. In two cases, aberrant bands in exon 2 of the VHL gene were demonstrated in the stromal cells by PCR-based single-strand conformation polymorphism analysis, and somatic missense mutations were successfully characterized in two of the sporadic hemangioblastomas by direct sequencing. The results suggest that allelic losses and mutations of the VHL tumor suppressor gene play a role in sporadic cerebellar hemangioblastoma tumorigenesis. Furthermore, because the genetic changes were detected in selectively procured stromal cell areas, the data provide strong evidence that the stromal cell represents a neoplastic component of hemangioblastoma.

Quantitative analysis of mitochondrial DNA deletion in paraffin embedded muscle tissues from patients with KSS and CPEO.

The percentage of common deletion of mitochondrial DNA (mtDNA) was determined quantitatively by a PCR-based, non-radioactive method in DNA extracted from formalin-fixed, paraffin-embedded skeletal muscle tissues from two patients with Kearns Sayre syndrome (KSS) and one with chronic progressive external ophthalmoplegia (CPEO). The method involved PCR cycle titration of wild-type and deleted mtDNA in parallel, staining of gel bands with the sensitive fluorescence dye SYBR Green I, and quantitation of intensity on a computer screen by the NIH image program. We determined 75% and 71% common deletion of mtDNA in the KSS patients and 35% in the CPEO patient.

Left-right asymmetries of the temporal speech areas of the human fetus.

Left-right asymmetries of the transverse temporal (Heschl) gyri and the temporal plane become recognizable by 31 weeks' gestation. The transverse temporal gyri are larger in number and extent on the right side in 54% of 207 serially sectioned fetal brains ranging in gestational age from 10 to 44 weeks, and the temporal plane is larger on the left side in those brains. There are two transverse temporal gyri on the left and a single right transverse temporal gyrus on the right in 18% of the brains. No asymmetry of number of transverse temporal gyri or extent of the temporal plane is apparent in 28%. These findings, which confirm those in adult brains, suggest that anatomical asymmetries for left hemispheral speech and language dominance may be established during the last trimester of fetal life.

Transient expression of transglutaminase C during prenatal development of human muscles.

Tissue transglutaminase (TGase C, TGase II) is known to participate in cellular processes during morphogenesis, differentiation, and development of various prenatal tissues and organs. The expression of TGase C during myoblast proliferation and attachment to external laminae was examined by immunohistochemical (IH) localization at 5-12 weeks of developmental stages of prenatal human muscle in 23 embryos. IH detection using a monospecific antibody to TGase C showed a prominent expression of TGase C in muscle cells as stage- and spatial-specific patterns during an early embryonal period. The myoblasts of intervertebral, tongue, and limb muscles, attached to adjacent cartilaginous skeletons or fibrous fascia, showed a pronounced expression of TGase C at 5-6, 6-7, and 7-8 weeks after fertilization, respectively. The most intense activity of TGase C was observed in some cardiac myoblasts infiltrating into endocardial mesenchyme at 6-7 weeks after fertilization. Although weak staining was detected until 14 weeks after fertilization, the level of TGase C expression in all muscles was significantly decreased after 6-7 weeks, with the exception that the smooth muscle cells of blood vessels and gastrointestinal tract showed diffusely intense staining of TGase C between 5 and 12 weeks after fertilization. Western blotting analysis of the cellular extracts of pooled samples showed a single strong band at 80 kD at 6 weeks after fertilization. This band became weaker after 8-10 weeks of prenatal development. These findings of transient expression of TGase C, which coincides with the development of myoblast anchoring and differentiation, suggest that TGase C plays a role in myoblast attachment to the extracellular laminae during the early embryonal period.

Bone pathologic correlation of multimodality imaging in Paget's disease.

The pagetic bones in the active phase of the disease with brisk lysis and sclerosis manifest intense tracer uptake on planar bone and SPECT images. Intense tracer uptake, however, can occur also in infections, dysplasias and metastases. Pinhole bone scintigraphy has been shown to portray specific diagnostic signs in a number of skeletal diseases. In an effort to identify useful bone scan signs, we prospectively carried out 99mTc-oxidronate pinhole bone scintigraphy of the skull, vertebrae, ribs, humerus, sacrum and ilium in two patients with Paget's disease of the bone. The pinhole bone scintigraphy findings correlated with radiographic, CT and MRI findings and in the vertebra with the pathological study. Interestingly enough, pinhole bone scintigraphy revealed intense tracer uptake preferentially in the bone cortex and the rim of the affected bones. Thus, the cranial inner table, humeral cortex and vertebral endplates and rims were the seats of characteristic tracer uptake, respectively creating a scintigraphic version of the radiographic "cotton wool" sign, "casket" sign and "picture frame" sign. The pagetic lesions in the sacrum and ilium also showed intense cortical and rim uptake. Correlation of pinhole bone scintigraphy with radiography, CT and MRI indicated that such cortical or rim uptake is characteristic of Paget's disease.

HLA-DR expression in human fetal thymocytes.

We analyzed the expression of MHC class I (W6/32) and class II (HLA-DR) antigens on human fetal and postnatal thymocytes by fluorescence-activated cell sorting. Less than 5% of prenatal thymocytes expressed HLA-DR before week 12 of gestation. However, the number of HLA-DR-positive cells significantly increased during the late second and third trimester of gestation, when greater than 50% of prenatal thymocytes expressed HLA-DR. Such high-level expressions of HLA-DR in fetal thymocytes were also demonstrated by Northern-blot analysis and immunohistochemistry. After birth, the percentage of HLA-DR-positive cells in thymocytes decreased gradually. A high-level expression of class I antigen was also observed in thymocytes from the early stages of gestation, but, in contrast to MHC class II, a majority of postnatal thymocytes maintained high levels of class I antigen after birth.

Overexpression of cyclin D1 and cdk4 in tumorigenesis of sporadic hepatoblastomas.

Abnormality of the cyclin D1/cdk4/p16INK4a/pRb pathway during tumorigenesis has recently been reported. Hepatoblastoma is a rare malignant liver tumor of childhood, but underlying abnormalities of cell-cycle regulating protein remain to be elucidated. The expression of cyclin D1, cdk4, p16 and retinoblastoma gene product (pRb) was studied by immunohistochemistry in 17 paraffin-embedded tissues consisting of both tumor and corresponding non-neoplastic tissues. Tumor tissues showed overexpression of cyclin D1 (13/17, 76%) and cdk4 (15/17, 88%). Eleven cases showed co-overexpression of both cyclin D1 and cdk4. No abnormal p16 or pRb expression was noted. In the group with a high score (+4) for cyclin D1 expression, a positive correlation with tumor recurrence was noted (P = 0.043). These data suggest that overexpressed cyclin D1 and cdk4 protein might play an important role in the tumorigenesis of hepatoblastoma and that in the group with high cyclin D1 expression, tumor recurrence may be more frequent.

Prevalence of Gs alpha mutations in Korean patients with pituitary adenomas.

The reported frequencies of Gs alpha mutations (gsp mutations) in growth hormone (GH)-secreting pituitary adenomas are variable (ranging from 4.4 to 43%), and the presence of these mutations in the other pituitary adenomas is still a matter of controversy. Previous clinical and biochemical analyses of patients with GH-secreting pituitary adenomas and gsp mutations produced conflicting results and did not demonstrate obvious characteristics. Therefore, we investigated the prevalence of gsp mutations in Korean patients with pituitary adenomas and elucidated the characteristics of these patients. Forty-four GH-secreting adenomas, 7 prolactin (PRL)-secreting adenomas and 32 clinically non-functioning adenomas were examined for the presence of point mutations in codon 201 and 227 of the Gs alpha gene using a nested PCR and direct sequencing of DNA extracted from fresh tissue or paraffin-embedded pituitary adenoma samples. Seven of the 44 GH-secreting pituitary adenomas had point mutations at codon 201 or 227; of these, five mutations were in codon 201 and two were in codon 227. In patients with gsp mutations, mean tumor size was significantly smaller than in patients without gsp mutations (15.9+/-8.7 mm vs. 24.9+/-14.9 mm, P<0.05). Age, sex, basal GH levels, GH response to oral glucose loading, GH response to octreotide and surgical outcome were not different in the two groups. One of the 32 clinically non-functioning pituitary adenomas had a point mutation at codon 201; none of the seven prolactinomas had these mutations. These results show that gsp mutations are not rare in Korean acromegalic patients and mean tumor size is significantly smaller in acromegalic patients with gsp mutations. Our results also confirm the low frequency of gsp mutations in clinically non-functioning pituitary adenomas and the absence of gsp mutations in prolactinoma.

The detection of p53 gene mutation using a microdissection technique in primary intracranial germ cell tumors.

Using a microdissection technique, the contribution of the p53 mutation to tumorigenesis and prognosis in each histological subtype of the intracranial germ cell tumors (GCTs) was evaluated. Nineteen patients had primary intracranial GCTs, including 4 germinomas (GEs), 4 teratomas (TEs), 1 mixed tumor of GE and TE, and 10 mixed GCTs containing non-germinomatous malignant germ cell tumors (NG-MGCTs). After microdissection of specific subtypes, genomic DNA was screened for mutations in exons 5-8 of the p53 gene, using the dideoxyfingerprinting (ddF) followed by direct DNA sequencing. The direct sequencing revealed a total of six mutations in PCR products derived from the five cases (26%) which showed mobility shifts in ddF. Among the six mutations detected, four were missense mutations and two were silent. Missense mutations of the p53 gene tended to occur more frequently in the NG-MGCT component than in the GE or TE components (3/15 vs. 1/12 vs. 0/13). The incidence of missense mutations was not different between the survivors (3/13) and the deceased (1/6). This study suggests the possible role of the p53 gene in the tumori-genesis of NG-MGCT. However, p53 gene mutation did not correlate with the prognosis of NG-MGCT.

Acute funisitis of preterm but not term placentas is associated with severe fetal inflammatory response.

Acute funisitis, whose basic pathologic feature is umbilical vasculitis, constitutes a type of fetal inflammatory response to intrauterine infection. In the present study, a comparative analysis was performed between the clinicopathologic profiles of acute funisitis in term and preterm placentas along with measurement of fetal plasma interleukin 6 (IL-6) levels by specific immunoassay to assess the different biologic implications for the fetus. Acute funisitis in preterm placentas showed a significantly higher incidence of umbilical arteritis (P <.000001), higher fetal plasma IL-6 level (P <.0001), and higher prevalence of major perinatal morbidities (P <.0001). To assess the possible variation in fetal cell response to infectious agents according to gestational age, amnion cells and placental villous tissues obtained at different gestational ages were treated with bacterial lipopolysaccharides, and the IL-6 level of the culture media was assayed. Amnion cells and placental villous tissues from preterm placenta showed a more pronounced cytokine response than those from term placenta. The findings of this study indicate that the clinicopathologic significance of acute funisitis in term placentas is different from that of preterm placentas. Furthermore, they indicate that the robust inflammatory response of the fetus associated with elevated fetal plasma IL-6 level may reflect the biologic needs of the premature fetus to escape from the hostile intrauterine environment.

The effect of regular salbutamol on lung function and bronchial responsiveness in patients with primary ciliary dyskinesia.

STUDY OBJECTIVE: There is growing evidence that regular beta(2)-agonist use in patients with asthma is associated with decreased airway caliber and increased bronchial responsiveness. The aim of this study was to determine whether regular treatment with beta(2)-agonists induces changes in lung function and bronchial responsiveness in patients with primary ciliary dyskinesia. DESIGN: A randomized, double-blind, placebo-controlled, crossover study. PATIENTS: Nineteen children with primary ciliary dyskinesia. INTERVENTIONS: Subjects received inhaled salbutamol or identical placebo (2 x 100 microg qid) for periods of 6 weeks with a wash-out period of 4 weeks. MEASUREMENTS AND RESULTS: FEV(1) was measured before and 3 weeks and 6 weeks after salbutamol or placebo treatment. High-dose methacholine inhalation tests were performed before and 6 weeks after each treatment. The provocative concentration of methacholine producing a 20% fall in FEV(1) (PC(20)) and maximal airway narrowing (MDeltaFFEV(1)) was measured. No significant change in FEV(1) was observed during the salbutamol or placebo periods. No significant differences in the parameters of bronchial responsiveness (PC(20) and MDeltaFFEV(1)) were noted as the result of either salbutamol or placebo treatment. CONCLUSION: Our data have shown that salbutamol, inhaled regularly for 6 weeks, did not cause either a decline in lung function or an increase in bronchial responsiveness in subjects with primary ciliary dyskinesia.

Sequential change of MR signal intensity of the brain after manganese administration in rabbits. Correlation with manganese concentration and histopathologic findings.

RATIONALE AND OBJECTIVES: High signal intensity in the basal ganglia on T1-weighted MR imaging has been reported in chronic manganese (Mn) poisoning. However, the exact meaning of the high signal intensity remains unclear: does it result from Mn itself, secondary pathologic changes of the brain tissue, or both? The goal of this study was to evaluate the sequential change of MR signal intensity and to correlate the MR intensity of the globus pallidus and the hypothalamus with the Mn concentration in the blood and the brain tissue, and with the histopathologic findings. METHODS: Ten milligrams per kilogram of Mn was administered once a week for 4 weeks to 14 rabbits. The rabbits in the control group (n = 2) were killed without Mn administration; those in group I (n = 4) were killed 1 day after the completion of Mn administration, those in group II (n = 4) were killed at 4 weeks, and those in group III (n = 6) were killed at 8 weeks. Sequential MR imaging, blood and tissue concentration measurement, and pathologic examination were performed. Sequential changes of the percent contrasts, contrast-to-noise ratios, and T1 relaxation times were analyzed with blood and tissue concentrations and histopathologic findings. RESULTS: The signal intensity of the basal ganglia on T1-weighted imaging was highest 1 day after cessation of Mn administration and sequentially washed out. The contrast, contrast-to-noise ratio, and T1 relaxation time showed significant correlations with blood concentration. Only the T1 relaxation time of the globus pallidus showed a significant correlation with tissue concentration. Histopathologic examination disclosed mild abnormalities in the globus pallidus, thalamus, and hypothalamus. CONCLUSIONS: The high signal intensity on T1-weighted MR imaging presumably indicates mainly the exposure marker of Mn, although mild pathologic findings were observed.

An unusual venous anomaly of the placenta.

The authors present an unusual vascular anomaly of the placenta. The placenta was very large, weighing 1,490 g. On the fetal surface, numerous dilated and tortuous vessels were observed on and under the chorionic membrane, of which three branches arose from a vein that was connected to the umbilical vein. One of them had a 5 x 2.5 cm aneurysmal dilatation, where three secondary branches arose. These venous channels were dilated and tortuous. The longest secondary branch was 133 cm in length and 1.2 cm in mean diameter and led into the placenta. Multiple, severely coiled or straight small branches arising from these vessels were also observed as vascular tangles. Some of these smaller vessels also led into the placenta. All abnormal vessels were veins. The umbilical cord was also normal except for a membranous insertion, and the placenta was unremarkable except for its large size.

Fetus-in-fetu: report of a case.

A case of fetus-in- fetu is reported. It occurred in an 8-week-old Korean boy who had been born by cesarean section due to abdominal distension. The fetus-in- fetu was connected to the superior mesenteric artery and consisted of two masses, apparently representing two portions of an acardiac monster. There was an amniotic membrane covering both masses, and the umbilical cord clearly was identifiable. One mass included the brain, eye, trachea, salivary glands, thyroid, pancreas, spleen, etc., while the other mass contained extremity bones, vertebrae, testis, adrenals, and intestinal loops. This is probably a case of separated fetus-in- fetu that showed unusually well-developed internal organs.

Reduced expression of CD99 and functional disturbance in anencephalic cortical thymocytes.

In a significant proportion of cases, anencephaly is associated with thymic enlargement, suggesting a possibility that anencephalic fetuses have a functional disturbance in thymocyte differentiation and development. In this report, we demonstrated that CD99 expression was consistently reduced in cortical thymocytes of all anencephalic fetuses. In addition, the CD99-dependent aggregation of immature cortical thymocytes was almost completely impaired and apoptosis of thymocytes was markedly reduced in several cases. These results are in agreement with previous findings that CD99 regulates the aggregation and apoptosis of various types of cells. These data strongly suggest that functional disturbance of thymocytes and thymic hyperplasia are related to the reduced expression of CD99 molecule in anencephalic fetuses.

Proliferation not apoptosis as a prognostic indicator in retinoblastoma.

The balance between proliferation and cell death is the major determinant of tumour growth. We analysed the proliferative and apoptotic indices (PI and AI, respectively) of 33 children with retinoblastoma. PI and AI were assessed by immunohistochemistry for Ki-67 antigen and TUNEL staining, respectively. The mean PI was 21.0+/-21.1%, and higher PI was associated with more advanced tumour stage (P<0.0001) and poor clinical outcome (P<0.05). Patients in whom amplified N-myc oncogene was found (n=6) determined by the multiplex polymerase chain reaction tended to have a higher PI (37.6+/-27.2%) than those without amplified N-myc (n=27; PI=17.3+/-18.1). A PI value of over 40% was clearly associated with an unfavourable prognosis. The AI, however, did not correlate with any of the other variables analysed. The findings suggest that proliferation, but not apoptosis, is of critical significance in retinoblastoma biology. PI, as determined by the Ki-67 antigen labelling index, seems to be a relevant histopathological parameter that can predict the clinical outcome of retinoblastoma.

Thymic epithelial tumor progression in an SV40T transgenic mouse model. Cortical thymoma-thymic carcinoma sequence.

There have been several reports that thymoma in human is a progressive disease, and that thymoma and thymic carcinoma form a continuum. We established a stable line of SV40T transgenic mice, which consistently produced thymic epithelial tumours progressing to thymic carcinoma within a predictable time span. Using this animal model and a morphological approach, thymic epithelial tumour progression was studied with reference to sequential changes at different time points in animals aged from 3 to 32 weeks. At all ages, SV40T was expressed in the nuclei of thymic epithelial cells; in these transgenic mice we observed the entire spectrum from cortical type thymoma to thymic carcinoma. Thymic size tended to increase with ageing in SV40T TG mice. While younger mice had predominantly cortical (organoid) or cortical thymoma, older mice had well-differentiated thymic carcinoma (WDTC) or poorly differentiated thymic carcinoma. When SV40T TG mice (248 line) reached a certain age, carcinoma of the thymus was present in all of them. Cortical-type thymoma became malignant within a predictable time span, suggesting a cortical thymoma-carcinoma sequence. When the mice were 9 weeks of age, the thymuses formed gross masses compatible with cortical thymoma. At 14 weeks of age, WDTC appeared against the background of cortical thymoma. Poorly differentiated thymic carcinoma was found after 15 weeks and affected all animals over 23 weeks of age. Most thymic carcinomas coexisted in varying proportions with cortical-type thymoma. Medullary thymomas did not develop in the mice, and no transition from medullary-type thymomas to thymic carcinomas was observed. In this SV40T transgenic mouse model, thymic carcinoma is clearly preceded by cortical-type thymoma. These transgenic mice may provide an interesting model for the progression from cortical thymoma to WDTC and/or high-grade carcinoma.

Hepatic capillariasis: first case report in the Republic of Korea.

We report a case of massive hepatic infection by Capillaria hepatica in a 14-month-old girl who presented with the symptom triad of persistent fever, hepatomegaly, and leukocytosis with eosinophilia. Twenty-five cases of human infection with this parasite, mostly in children, have been reported in the literature. This is the first case of hepatic capillariasis reported in the Republic of Korea. The diagnosis was made by needle biopsy of the liver. Scanning electron microscopic examination of the biopsy specimen was also performed. Thiabendazole therapy was initiated and the patient developed liver disease-related IgA nephropathy during the therapy. The literature dealing with proven cases of infection with C. hepatica is briefly reviewed.

Characterization of a mitochondrial DNA deletion in patients with mitochondrial myopathy.

Kearns-Sayre syndrome (KSS) is the most commonly diagnosed mitochondrial myopathy and it produces severe neuromuscular symptoms. The most frequent cause is a mitochondrial DNA (mtDNA) deletion (common deletion) that removes a 4,977-bp segment of DNA that includes several genes encoding respiratory chain subunits. In this disorder, the population of deleted mtDNA molecules coexists with a population of normal, wild-type, full-length mtDNAs. We have performed a morphological study and a 3-primer polymerase chain reaction (PCR) on paraffin-embedded muscle tissues from two Korean KSS patients. Our results show that these patients have the essential clinical manifestations of KSS, as well as morphological evidence of mitochondrial myopathy. PCR analysis revealed the existence of two co-amplified fragments, the wild-type fragment having 152 bp and the deleted fragment having the 123 bp characteristic of common deletion. This study may provide valuable information for the molecular diagnosis of KSS in Koreans.

Morphological study of surgically induced open neural tube defect in old (14 and 21 days) chick embryos.

As an experimental model for the research of open neural tube defect (NTD), the surgical model has several advantages over others, in spite of the fact that the pathogenetic mechanism is not compatible with the human intrauterine events. To make reproducible NTDs by surgery and to compare the surgically induced lesions with the human myeloschisis morphologically, we opened the neural tube for a length of 9-11 somites in Hamburger and Hamilton stage 16-19 chick embryos. Embryos which survived until the late in ovo life (total age 14 and 21 days) showed relatively reproducible open NTDs. Morphologically they are similar to human myeloschisis. This study suggests that the surgical model can be suitable for studies of open NTDs.