Excessive Subchorionic Fibrinoid Deposition as a Component of Massive Perivillous Fibrin Deposition: A Case With Maternal Immune Thrombocytopenia.

Maternal floor infarction (MFI) and massive perivillous fibrin deposition (MPFD) are overlapping placental disorders of unknown etiology, associated with adverse obstetric outcome, and a significant risk of recurrence. We describe a 31-year-old mother with asymptomatic thrombocytopenia throughout pregnancy and a positive lupus anticoagulant. She delivered a normal female neonate at term, whose weight was small for gestational age, with a placenta weighing less than the 10th percentile. Placental examination showed MPFD together with excessive subchorionic fibrinoid deposition. The placenta showed diffuse C4d deposition and an immune-mediated reaction was postulated for the pathogenesis of the placental changes. We suggest that excessive subchorionic fibrinoid deposition may be part of the morphologic spectrum of MFI/MPFD.

Classification of stillbirth by the International Classification of Diseases for Perinatal Mortality using a sequential approach: A 20-year retrospective study from Thailand.

AIM: The International Classification of Diseases for Perinatal Mortality (ICD-PM) is a system for recording causes of perinatal death. In this system, placental pathology is considered a "maternal condition" and this category does not cover the spectrum of placental pathology that can impact on perinatal death. The aim of the study was to apply a wider spectrum of placental pathology as a separate parameter for classifying death in the ICD-PM. METHODS: All autopsy reports at a single institution over a 20-year period (2001-2020) were reviewed. Causes of stillbirth were analyzed in a sequential manner: step 1, clinical history and laboratory results; step 2, placenta; and step 3, autopsy; and classified at each step according to the ICD-PM. RESULTS: The review identified 330 cases, including 126 antepartum and 204 intrapartum deaths. Step 1 identified a cause in 176 (86%) intrapartum deaths and 64 (51%) antepartum deaths. The addition of placental pathology (step 2) changed the cause of death in 12% of cases, with causes now identified in 190 (93%) intrapartum and 89 (71%) antepartum deaths. Adding step 3 did not identify any additional causes of death. CONCLUSION: The accuracy of the ICD-PM classification is dependent on the data available. Placental pathology made a significant difference in assigning causes of death in our series, stressing the importance of placental examination. Determination of the cause of death based on clinical history and laboratory data alone may be inaccurate, and less useful for comparative studies and planning prenatal care.

Pediatric fibromyxoid soft tissue tumor with PLAG1 fusion: A novel entity?

The classification of undifferentiated soft tissue tumors continues to evolve with the expanded application of molecular analysis in clinical practice. We report three cases of a unique soft tissue tumor in young children (5 months to 2 years old) displaying a purely fibromyxoid histology, with positive staining for desmin and CD34. In two cases, RNA sequencing detected a YWHAZ-PLAG1 gene fusion, while in the third case, a previously unreported EEF1A1-PLAG1 fusion was identified. PLAG1 fusions have been reported in several pathologic entities including pleomorphic adenoma, myoepithelial tumors of skin and soft tissue, and lipoblastoma, the latter occurring preferentially in young children. In these tumors, expression of a full length PLAG1 protein comes under the control of the constitutively active promoter of the partner gene in the fusion, and the current cases conform to that model. Overexpression of PLAG1 was confirmed by diffusely positive immunostaining for PLAG1 in all three cases. Our findings raise the possibility of a novel fibromyxoid neoplasm in childhood associated with these rare PLAG1 fusion variants. The only other report of a PLAG1-YWHAZ fusion occurred in a pediatric tumor diagnosed as a "fibroblastic lipoblastoma." This finding raises the possibility of a relationship with our three cases, even though our cases lacked any fat component. Further studies with regard to a shared pathogenesis are required.

Adult-onset neuroblastoma: Report of seven cases with molecular genetic characterization.

Whereas neuroblastoma is the most common extracranial solid tumor of childhood, less than 5% of cases occur in adults. Pediatric neuroblastoma shows marked heterogeneity of histology and molecular biology. Information about this tumor in adults is limited, especially regarding molecular biology. We report a series of nine neuroblastoma cases diagnosed in adulthood (18 to 40 years old) with molecular biologic characterization in seven. All tumors were Schwannian stroma-poor, and mostly poorly differentiated. Tumors expressed neural markers including PHOX2B, NB84, synaptophysin, chromogranin, CD56, neuron-specific enolase, and PGP9.5. Two out of six cases expressed ALK and one had the F1174 L mutation reported in childhood neuroblastoma. Fluorescent in situ hybridization (FISH) revealed MYCN amplification in 2/7 cases, chromosome 1p deletion in 1/5 cases and 17q gain in 4/4 cases. One in five cases showed loss of ATRX expression by immunohistochemistry and alternate lengthening of telomeres by FISH. Zero out of five cases showed rearrangement of the TERT gene by FISH, but one case showed high level amplification. In conclusion, the morphology and immunophenotype of adult-onset neuroblastoma are similar to pediatric cases although less differentiated than some childhood tumors. Similarly, molecular genetic alterations in adult-onset neuroblastoma are not unique to this age group. However, 80% of cases tested showed genetic changes that would promote maintenance of telomeres, which is a molecular marker of high risk cases. This may help explain the poor response in adults to pediatric treatment protocols. Additional studies to characterize the biology of this tumor in the adult age group will facilitate the design of more personalized therapeutic approaches.

Electron Microscopy Can Still Have a Role in the Diagnosis of Selected Inborn Errors of Metabolism.

Many anatomic pathology laboratories no longer have electron microscopy facilities. A retrospective review of autopsies was performed to identify cases of inborn errors of metabolism (IEM) and determine the contribution of electron microscopy in making the diagnosis in those cases. Over a period of 17 years, there were 900 perinatal and pediatric autopsies. There were 7 cases (1%) of IEM, including 4 cases of Pompe disease, 1 case of I-cell disease, 1 case of bile acid synthesis defect, and 1 case of mitochondrial disease (Leigh syndrome). Electron microscopy was important in the diagnosis of I-cell disease and Pompe disease in our series. This technique enabled a prenatal diagnosis to be made from a chorionic villus biopsy in 2 cases with a positive family history. In less developed countries where upfront genetic testing may be too expensive and may need international referral, electron microscopy can still be useful for diagnosis of IEM, providing an affordable alternative with a more rapid turnaround time compared to gene mutation analysis or enzyme assay. Results can be used both for patient management and as a screen for which cases might benefit from genetic testing.

Anaplastic sarcomas of the kidney are characterized by DICER1 mutations.

Anaplastic sarcoma of the kidney is a rare tumor (≤25 reported cases) characterized by the presence of cysts, and solid areas composed of bundles of undifferentiated spindle cells, showing marked cellular anaplasia (usually accompanied by TP53 overexpression). These tumors often feature prominent areas of cartilage or chondroid material. Germline mutations in DICER1, encoding the microRNA (miRNA) processor DICER1, cause an eponymous syndrome. Recent reports suggest that anaplastic sarcoma of the kidney should be included in DICER1 syndrome as germline DICER1 mutations are associated with the occurrence of such tumors. Therefore, we sought to determine the following: (1) what proportion of anaplastic sarcoma of the kidney have DICER1 mutations; (2) whether the identified mutations affect both alleles of DICER1 (ie, are biallelic); (3) whether somatic missense mutations in the DICER1 RNase IIIb domain impact miRNA generation; and (4) whether TP53 alteration always occurs in these tumors. DICER1 mutations were evaluated by Sanger sequencing and next-generation sequencing in nine tumor/normal pairs. Impact of DICER1 mutations on miRNA generation was evaluated via an in vitro DICER1 cleavage assay. TP53 status was assessed by immunohistochemistry and next-generation sequencing. Eight of the nine cases had at least one RNase IIIb DICER1 mutation that impacted the generation of miRNAs. There were six tumors with truncating DICER1 mutations and in four of them, the mutation found in the tumor was also detected in adjacent normal tissue, and therefore was likely to be either mosaic or germline in origin. Analysis of mutation phase revealed that two of three tumors had biallelic DICER1 mutations. Six of nine anaplastic sarcomas of the kidney had aberrant TP53 immunohistochemisty with damaging TP53 mutations identified in three cases. Taken together, these data suggest that the great majority of anaplastic sarcomas of the kidney have DICER1 mutations and confirm that these tumors are part of the DICER1 syndrome.

Heterogeneity of MYCN amplification in neuroblastoma at diagnosis, treatment, relapse, and metastasis.

Amplification of the MYCN gene in neuroblastoma is associated with a poor prognosis and is considered to remain unchanged in post-treatment specimens and metastases. While heterogeneity of MYCN copy number in tumor cells has been reported, serial samples have only been studied in a limited way, and the biologic relevance of this finding is not well understood. We used in situ hybridization on paraffin sections of 102 specimens from 30 patients with MYCN-amplified neuroblastoma to determine MYCN copy number in the primary tumor, pre- and post-treatment, and in metastatic samples. Nineteen cases (63%) showed diffuse MYCN amplification in all samples tested. Nine cases (30%) showed a reduction in MYCN copy number: five cases with diffuse amplification subsequently showed focal amplification, one case with diffuse MYCN amplification showed MYCN gain after treatment, and three focally amplified cases were non-amplified in later specimens. In two cases (7%), focal amplification became diffuse in subsequent samples. Histology was not predictive of the temporal or spatial pattern of MYCN amplification for a particular tumor. If extent of amplification (focal vs. diffuse) is not considered, 26/30 (87%) of cases were consistently MYCN-amplified. However, our data suggest that MYCN status can be heterogeneous between tumor sites, during tumor progression or following treatment, challenging the notion that MYCN copy number does not change for a particular neuroblastoma. Assessing the biologic significance of MYCN heterogeneity will require larger studies of clinically annotated tumor samples, and will depend on interpreting heterogeneity in MYCN status in combination with other genetic changes. © 2016 Wiley Periodicals, Inc.

The neuroblastoma and ganglion components of nodular ganglioneuroblastoma are genetically similar: evidence against separate clonal origins.

Nodular ganglioneuroblastoma is characterized by a macroscopic nodule of neuroblastoma within a ganglioneuromatous component. These two components have been considered to originate from separate clones, with the neuroblastoma clone accounting for the clinical behavior of nodular ganglioneuroblastoma. In order to investigate the clonal origin of the cellular components (neuroblasts, ganglion cells, and Schwann cells) of nodular ganglioneuroblastoma, paraffin-embedded tumor samples from eight cases were analyzed by single nucleotide polymorphism array and in situ hybridization. DNA was extracted separately from neuroblastomatous and ganglioneuromatous areas. By in situ hybridization, MYCN gain (4-10 gene copies/nucleus) was detected in 7/8 neuroblastoma samples. In ganglioneuromatous regions, gains were also detected in ganglion cells but not in Schwann cells. Single-nucleotide polymorphism array studies identified chromosome losses (11q and 14q) and gains (12, 13q, 17q and 18q) in the neuroblastoma component, whereas the ganglioneuromatous component showed fewer or no genetic alterations. There were no unique copy number changes distinguishing nodular ganglioneuroblastoma from other subtypes of neuroblastoma. By in situ hybridization, ganglion cells but not Schwann cells showed the same alterations detected in neuroblasts. Thus, neuroblasts and ganglion cells in nodular ganglioneuroblastoma are genetically related and may arise from the same clone. In contrast, the Schwann cells have a different origin and may be derived from a non-neoplastic neural crest precursor. Our results suggest that the clinical behavior of nodular ganglioneuroblastoma cannot be explained by the presence of separate clones with distinct genetic signatures.

Nestin is not essential for development of the CNS but required for dispersion of acetylcholine receptor clusters at the area of neuromuscular junctions.

Nestin is expressed in many different progenitors during development including those of the CNS, heart, skeletal muscle, and kidney. The adult expression is mainly restricted to the subependymal zone and dentate gyrus of the brain, the neuromuscular junction, and renal podocytes. In addition, this intermediate filament protein has served as a marker of neural stem/progenitor cells for close to 20 years. Therefore it is surprising that its function in development and adult physiology is still poorly understood. Here we report that nestin deficiency is compatible with normal development of the CNS. The mutant mice, however, show impaired motor coordination. Furthermore, we found that the number of acetylcholine receptor clusters, the nerve length, and the endplate bandwidth are significantly increased in neuromuscular junction area of nestin-deficient mice. This is similar to the phenotype described for deficiency of cyclin-dependent kinase 5 (Cdk5), a candidate downstream affecter of nestin. Moreover, we demonstrate that nestin deficiency can rescue maintenance of acetylcholine receptor clusters in the absence of agrin, similar to Cdk5/agrin double knock-outs, suggesting that the observed nestin deficiency phenotype is the consequence of aberrant Cdk5 activity.

Natural course of low risk neuroblastoma.

BACKGROUND: Neuroblastoma is characterized by heterogeneity of histology, biology, and clinical behavior. Most epidemiology studies are based on Western and Japanese populations; there are very few studies on neuroblastoma from Southeast Asia. PROCEDURE: Cases of Thai children with neuroblastoma were retrospectively reviewed to determine if the epidemiology of the disease differs from Western populations. Sixty-two cases were assembled from two pathology centers in Bangkok. Histologic prognostic category and MYCN copy number were determined. RESULTS: The median age at diagnosis was 2.9 years. Only 11% of cases presented at less than 1 year of age and 12% cases had low stage disease (1, 2, and 4S). The majority of tumors had unfavorable histology (48/62); this was at least partly due to the higher age at diagnosis for most patients. MYCN amplification was detected in 18/52 (35%) tumors, all in stage 3 or 4 tumors. We assigned patients to high, intermediate and low risk categories using the Children's Oncology Group risk stratification criteria. In contrast to Western studies, the majority of cases (50/59 or 85%) in our series had high risk disease. CONCLUSIONS: Since there is no evidence to date that the biology of neuroblastoma varies by geographic region, the paucity of low risk cases in our study may reflect spontaneous resolution/differentiation of tumors that are not clinically detected. Moreover, a delay in diagnosis of intermediate risk cases could result in higher tumor burden at the time of diagnosis, increasing the proportion of high risk cases observed.

Primary intracranial Ewing sarcoma with an unusually aggressive course: a case report and review of the literature.

The occurrence of Ewing sarcoma-peripheral primitive neuroectodermal tumor as a primary intracranial tumor is very rare, with only 29 cases reported in the literature, 19 of which have included molecular studies. We present the clinical, radiologic and pathologic findings of an intracranial Ewing sarcoma in a 22-year-old woman arising from the dura over the right frontal convexity. The patient underwent craniotomy with gross total excision of the tumor. The tumor showed atypical histology and the diagnosis was confirmed by detection of a rearrangement of the EWSR1 gene by fluorescent in situ hybridization and identification of the diagnostic t(11;22)(q24;q12) translocation by reverse transcription-polymerase chain reaction. Additional features were detected in this tumor that are known to be associated with an unfavorable prognosis, including loss of p16 expression and gains of chromosomes 1q and 12. The patient experienced the most rapid downhill course reported to date for intracranial Ewing sarcoma, developing multiple extracranial metastases at 2 months and dying 6 months after the initial operation.

PRAME protein expression in DICER1-related tumours.

DICER1 syndrome is an autosomal dominant tumour predisposition syndrome usually affecting persons under 30 years of age. Many of the associated benign and malignant lesions occur almost exclusively in DICER1 syndrome. One such tumour, pituitary blastoma (pitB), overexpresses PRAME 500x above control levels. PRAME (PReferentially expressed Antigen in MElanoma) is expressed in malignancies that are not DICER1-related (e.g. melanoma). To address whether PRAME expression is part of the DICER1 phenotype, or simply a feature of pitB, a series of 75 DICER1-mutated specimens and 33 non-mutated specimens was surveyed using immunohistochemistry for PRAME, together with EZH2, which complexes with PRAME. In DICER1-mutated specimens, positive staining for PRAME was only seen in malignant tumours; 7 of 11 histological types and 34/62 individual tumours were positive, while non-tumourous lesions were always negative. Pleuropulmonary blastoma (PPB) showed a continuum in staining, with type I lesions being PRAME negative (n = 7) but all type II and type III lesions PRAME positive (n = 7). Similarly, cystic nephroma (CN) was negative (n = 8), with anaplastic sarcoma of the kidney being positive (n = 2). However, one atypical CN with mesenchymal cell proliferation was PRAME-positive. Embryonal rhabdomyosarcoma (RMS) with DICER1 pathogenic variants (PVs) was positive for PRAME (5/6), but the same tumour type without DICER1 PVs was also positive (9/15). Staining for EZH2 corresponded to that seen with PRAME, validating the latter. This study leads us to conclude that (1) PRAME expression occurs in two-thirds of DICER1-related malignancies; (2) PRAME may be a marker for the progression that certain DICER1-related lesions are thought to undergo, such as PPB and CN; and (3) PRAME expression in some tumours, such as RMS, appears to be an intrinsic feature of the tumour, rather than specifically related to DICER1 PVs. Therapy directed against PRAME may offer novel treatment options in patients with the DICER1 syndrome.

Paediatric BCOR-associated sarcomas with a novel long spliced internal tandem duplication of BCOR exon 15.

Clear cell sarcoma of the kidney (CCSK) and primitive myxoid mesenchymal tumour of infancy (PMMTI) are paediatric sarcomas that most commonly harbour internal tandem duplications (ITDs) of exon 15 of the BCOR gene, in the range of 87-114 base pairs (bp). Some cases, instead, have BCOR-CCNB3 or YWHAE-NUTM2 gene fusions. About 10% of cases lack any of these genetic alterations when tested by standard methods. Two cases of CCSK and one PMMTI lacking the aforementioned mutations were analysed using Archer FusionPlex technology. Two related BCOR exon 15 RNA transcripts with ITDs of lengths 388 and 96 bp were detected in each case; only the 388 bp transcript was identified when genomic DNA was sequenced. In silico analysis of this transcript revealed acceptor and donor splice sites indicating that, at the RNA level, the 388-bp transcript was likely spliced to form the 96-bp transcript. The results were confirmed by Sanger sequencing using primers targeting the ITD breakpoint. This novel and unusually long ITD segment is difficult to identify by DNA sequencing using typical primer design strategies flanking entire duplicated segments because it exceeds the typical read lengths of most sequencing platforms as well as the usual fragment lengths obtained from formalin-fixed paraffin-embedded material. As diagnosis of CCSK and PMMTI may be challenging by morphology and immunohistochemistry alone, it is important to identify mutations in these cases. Knowledge of this novel BCOR ITD is important in relation to primer design for detection by sequencing, and using RNA versus DNA for sequencing.

Simplified Molecular Subtyping of Medulloblastoma for Reduced Cost and Improved Turnaround Time.

Molecular subtyping of medulloblastoma (MB) has become increasingly important for prognosis and management. Typically this involves detailed molecular genetic testing which may not be available in all centers. The purpose of the present study was to find a simplified approach to assign molecular subtypes of MB for routine use in centers with more limited resources. The molecular subtypes of MBs from 32 Thai patients, aged 0.5 to 35 years, were first determined by NanoString. These results were then compared with those obtained using a combination of limited immunohistochemistry (IHC) (ß-catenin, GAB-1, YAP-1, p75-NGFR, OTX2) and CTNNTB exon 3 mutation analysis. By NanoString assay, there were 6 MBs (19%) in the wingless (WNT) group, 8 (25%) in the sonic hedgehog (SHH) group, 7 (22%) in group 3, and 11 (34%) in group 4. Although ß-catenin immunostaining missed 4/6 WNT MBs, CTNNTB mutation analysis confirmed all WNT MB cases with amplifiable DNA. The IHC panel correctly assigned all the other molecular subtypes, except for 1 MB in group 4. Thus, our protocol was able to correctly categorized 31/32 cases or 97% of cases. Our study is the first to report molecular subtypes of MB in Southeast Asia. We found that molecular subgroups of MBs can be reliably assigned using a limited IHC panel of ß-catenin, GAB-1, YAP-1, p75-NGFR, OTX2, together with CTNNTB exon 3 mutation analysis. This simplified approach incurs lower cost and faster turnaround time compared with more elaborate molecular methodologies and should be beneficial to centers with reduced laboratory resources.

DICER1 screening in 15 paediatric paratesticular sarcomas unveils an unusual DICER1-associated sarcoma.

Individuals with DICER1 syndrome, a genetic disorder caused by pathogenic germline variants in DICER1, are at increased risk of developing a wide array of predominantly childhood onset conditions, including genitourinary sarcomas. However, data on DICER1 involvement in paratesticular sarcomas have not been published. Herein, we analyse a series of 15 paediatric paratesticular sarcomas and describe in detail the case of a male infant with a paratesticular myxoid tumour, considered to be a low-grade sarcoma, who also manifested a cystic nephroma, a classic DICER1 syndrome phenotype. He harboured a pathogenic germline DICER1 variant and different somatic hot-spot mutations in each tumour. The paratesticular tumour showed strong and diffuse expression for WT1 and CD10, an unusual immunophenotype in paediatric sarcomas, but typical of tumours of Müllerian origin. The tumour was postulated to arise from the appendix testis, a Müllerian remnant located in the paratestis. Such an origin would be analogous to other DICER1-associated non-epithelial gynaecological tumours, thought to arise from Müllerian derivatives. These findings point towards a key role of DICER1 in Müllerian-derived structures. Supporting this hypothesis is the fact that the other paratesticular sarcomas from the series were either negative or focally positive for WT1 and for CD10, and none had any DICER1 mutations. In summary, we present the first case of a paratesticular sarcoma associated with DICER1 syndrome, emphasising that paratesticular tumours with an unusual histological appearance may suggest an underlying DICER1 mutation, especially in the presence of a personal or family history of DICER1-associated disease. In this context, DICER1 mutation testing could lead to changes in clinical care including implementation of cancer care surveillance strategies.

Cyclooxygenase-2 expression in squamous cell carcinoma of the uterine cervix is associated with lymph node metastasis.

OBJECTIVES: Previous studies have indicated that cyclooxygenase-2 (COX-2) activity is related to the development and progression of cervical cancer. In this study, we evaluated the association between COX-2 expression and specific clinicopathologic features in surgically-treated squamous cell carcinoma of the uterine cervix. METHODS: Immunohistochemical staining for COX-2 was performed on 196 cases of stage IB-IIA cervical squamous cell carcinoma. Results were correlated with the clinicopathologic features and disease-free survival using statistical analysis. RESULTS: Expression of COX-2 was detected in 48.5% of cases. COX-2 expression was significantly associated with lymph node metastasis (p=0.045) but lacked significant correlation with tumour stage, size, histologic grade, deep stromal invasion, lymphovascular space invasion, and parametrial involvement. In multivariate analysis, only parametrial involvement and lymphovascular space invasion (LVSI) were independent predictors for lymph node metastasis (p=0.001 and 0.007, respectively). COX-2 expression was not associated with lymph node metastasis in the absence of parametrial involvement or LVSI. In the cases with LVSI, COX-2 expression was significantly associated with lymph node metastasis (p=0.03), although with marginal significance (p=0.068) in the multivariate analysis. COX-2 expression was not associated with a decrease in disease-free survival for patients overall (p=0.977). However, in patients who did not receive adjuvant treatment, COX-2 expression was significantly associated with decreased disease-free survival (p=0.008) and was a significant predictor of recurrence (p=0.014). CONCLUSIONS: In this study, COX-2 expression was associated with lymph node metastasis in cervical squamous cell carcinoma, but this was linked to the presence of LVSI or parametrial involvement. This suggests that COX-2 expression may enhance lymph node metastasis after LVSI occurs. If so, immunohistochemical staining for COX-2 may provide additional prognostic information in LVSI-positive cases, in particular in patients who do not receive postoperative adjuvant treatment.

Fibrolamellar carcinoma presenting as a pancreatic mass: case report and review of the literature.

Fibrolamellar carcinoma is a subtype of hepatocellular carcinoma with distinct clinicopathologic features including presentation at a younger age. Although early studies suggested that fibrolamellar carcinoma had a better prognosis than conventional hepatocellular carcinoma, most later studies have found no difference. Patients often have lymph node metastases at presentation in addition to the hepatic primary. We describe an unusual case in a Thai boy who presented with a pancreatic mass that was clinically suspected to be a primary pancreatic tumor, but on biopsy was found to be metastatic fibrolamellar carcinoma. To our knowledge, this manner of presentation has not been previously reported for fibrolamellar carcinoma, nor has metastatic spread to the pancreas.

J1-31 protein expression in astrocytes and astrocytomas.

J1-31 is one of the astrocytic proteins, the expression of which has not been evaluated in astrocytomas. In the present study, we studied the expression of J1-31 protein in astrocytes and astrocytomas in comparison with GFAP, p53 and Ki-67. Materials consisted of formalin-fixed paraffin-embedded tissue specimens that included five cases of normal brain, 17 of gliosis, 15 of pilocytic astrocytoma (WHO grade I), 26 of low-grade diffuse astrocytoma (WHO grade II), four of anaplastic astrocytoma (WHO grade III), and eight of glioblastoma (WHO grade IV). GFAP was highly expressed in all specimens examined. The anti-J1-31 antibody exhibited strong cytoplasmic staining of reactive gliosis in 17/17 (100%) cases with a higher intensity of staining than that observed in the adjacent normal astrocytes. The antibody showed reactivity with tumor cells in 12/15 (80%) cases of pilocytic astrocytoma, although intensity of staining was generally weaker and more focal than observed in reactive gliosis. J1-31-positive tumor cells were detected in only 9/26 (35%) cases of the low-grade diffuse astrocytoma and none of the cases of anaplastic astrocytoma and glioblastoma. Increasing Ki-67 indices paralleled advancing tumor grades. p53 protein was expressed more commonly in infiltrating astrocytomas compared to pilocytic astrocytoma. In conclusion, down-regulation of J1-31 expression correlates with advancing grade of astrocytomas. The result suggests this protein plays some role in astrocytes that is progressively lost in malignant progression. The anti-J1-31 antibody may help further our understanding of astrocytes in disease and may be useful as an aid in the pathologic diagnosis of astrocytic lesions.

Podocytes contribute to the formation of glomerular crescents.

The cellular composition of crescents in glomerular disease is controversial. The role of podocytes in crescent formation has been especially difficult to study because podocytes typically lose their characteristic terminally differentiated phenotype under disease conditions, making them difficult to identify. We reasoned that the intermediate filament protein nestin, a marker of progenitor cells that has recently been identified in podocytes, may allow the investigation of podocyte involvement in glomerular crescents. In a series of 35 biopsies with crescentic glomerular disease, all showed nestin-positive cells in the crescents, ranging in number from occasional to approximately 50% of crescent cells. Other podocyte markers, such as podocin and WT1, failed to identify cells in crescents, and no contribution by endothelial or myogenic cells was noted. CD68-positive cells were observed in 80% of cases but were never as numerous as the nestin-positive cells. Nestin and CD68 were not coexpressed by the same cells, providing no evidence of trans-differentiation of podocytes into a macrophage phenotype. Keratin-positive cells were found in crescents in 51% of cases, but only as occasional cells. Up to one third of crescent cells were cycling in 48% of biopsies, and double immunostaining identified these cells as a mixture of nestin-positive cells and "null" cells (negative for nestin, CD68, and keratin). In addition to our observations in human disease, we also identified nestin-positive proliferating podocytes in the crescents of 2 mouse models of crescentic glomerulonephritis. We conclude that podocytes play a role in the formation of glomerular crescents.