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Clustering of features and genetics of von Hippel-Lindau syndrome SiR,-In response to our report (May 4, p 1052) on the clustering of clinical features in von Hippel-Lindau syndrome (HLS), Dr Filling-Katz and Dr Choyke, Dr Maher, and Professor Langenbeck (June 15, pp 1477-79) raise several interesting issues. Maher reports a heterozygote prevalence of 1 in 53 000 for HLS in East Anglia.This figure is in good agreement with our prevalence of 1 in 38 951, indicating that the Freiburg kindreds do not represent an epidemiological cluster of HLS. Apparently, there are occasional families presenting with both renal cell carcinoma and phaeochromocytoma. However, the great majority of recorded HLS kindreds show only one of these manifestations. An especially striking example is reported by Glenn et al.2 They observed an HLS family with 47 affected members, 57% of whom had phaeochromocytomas. Renal and pancreatic lesions were not detectable in any of these patients. In the same study most families presenting with renal and pancreatic features did not exhibit
phaeochromocytomas. Maher points out that
Department of Pathology, University of Zurich Zurich, Switzerland 1. Maher
Germany
HARTMUT P. H. NEUMANN
OTMAR D. WIESTLER
ER, Iselius L, Yates JRW, et al. Von Hippel-Lindau disease: a genetic study.
J Med Genet (m press). GM, Daniel LN, Choyke P, et al. von Hippel-Lindau disease: distinct phenotypes suggest more than one mutant allele at the VHL locus. Hum Genet (in press). 3. Neumann HPH. Basic criteria for clinical diagnosis and genetic counselling in von Hippel-Lindau syndrome. J Vasc Dis 1987; 16: 220-26. 4. Neumann HPH, Eggert HR, Weigel MD, et al. Hemangioblastoma of the central nervous system: a 10-year study with special reference to von Hippel-Lindau syndrome. J Neurosurg 1989; 70: 24-30. 5. Neumann HPH, Dinkel E, Brambs H, et al. Pancreatic lesions in the von Hippel-Lindau syndrome. Gastroenterology (in press) 2. Glenn
Fetal
DMD
family.
Arrow indicates carrier at risk.
renal carcinomas are frequently not detectable before the age of 44 (mean age of diagnosis in his study) and may, therefore, be missed in younger HLS patients. This cannot account for our results, since 12/28 of our patients from kindreds without renal lesions have passed the age of 44. Maher expresses concern that our focus on clustered manifestation of HLS may distract the attention for other features of the syndrome. All patients and potential gene carriers underwent a broad screening examination. 3,4 Filling-Katz and Choyke describe families with additional islet cell tumours and pancreatic adenocarcinoma as yet another example of a cluster of HLS features. However, it remains to be determined if these manifestations, which are most unusual in our HLS patients,s really represent manifestations of the syndrome. Several genetic models could account for the distinct patterns of features in HLS. We agree that, besides a complex genetic locus with several transcriptional units or cooperating genes on different chromosomes, allelic heterogeneity of the HLS gene on chromosome 3p is another potential genetic mechanism. Only a detailed molecular analysis in representative kindreds of the still elusive HLS gene will allow us to identify the genetic basis of these heterogeneous features. Department of Medicine, Albert Ludwig’s University, 7800 Freiburg im Breisgau,
Fig 1-Pedigree of Dutch
recombination distal to the DMD gene betwee H and F. However, the woman seeking advice had inherited an X-chromosome with a
potentially intragenic recombination between the proximal intragenic probe L and the first informative distal probe F. Although she had normal creatine kinase values, her carrier risk was high, possibly 20%. First trimester prenatal diagnosis on chorionic villi of her male pregnancy (III-2) showed the at-risk haplotype and the parents opted for termination. Muscle tissue of the fetus was studied immunohistochemically for dystrophin expression. We used monoclonal antibodies against the 30 kDa segment of dystrophin2 (aminoacids 1181-1388),3a59 kDa segment (aminoacids 17502248),4or the carboxy-terminal 17 aminoacids. All three showed clear staining of dystrophin in parts of the plasma membrane and at the distal ends of the myotubes5 (fig 2), an immunohistochemical pattern indistinguishable from that routinely found in normal fetuses. In affected fetuses, dystrophin cannot be detected with antibodies reactive with the distal part of the protein and sometimes not with
central antibodies either.6 Thus, this fetus was not affected and the recombination event in 11-2 has not included the DMD mutation. Her carrier risk was reduced to the population risk. Bieber et aF demonstrated by negative western blot testing that a 23-week-old DMD at-risk fetus was affected, confirming the DNA diagnosis and raising the mother’s carrier risk to 100%. We show here that even in younger fetuses sensitive dystrophin immunohistology permits genetic counselling. This will be especially valuable in DMD families where no mutation is found and the DNA markers are uninformative or show intragenic recombinations.
dystrophin to diagnose carrier status
SIR,-In a Dutch family with Duchenne muscular dystrophy (DMD) we have excluded carriership for a woman at risk (11-2, fig 1) by immunohistochemical analysis of her 12-week-old fetus after abortion for increased DMD risk. In this family, DNA analysis was uninformative. Two relatives (11-3,11-5) were affected with DMD. cDNA and polymerase chain reaction studies failed to show a mutation in the DMD gene, so the at-risk chromosome had to be identified by haplotype analysis.The informative markers used (H, F, L, C, and T) are identified in fig 1. Two non-carrier daughters were identified; 11-4 had the unaffected haplotype and 11-6 had a
Fig 2-lmmunohistological studies of fetal muscle. Tissue fixed and stained" and sections
were
carboxy-terminal monoclonal antibody.
was
immunostained with
Clustering of features and genetics of von Hippel-Lindau syndrome SiR,-In response to our report (May 4, p 1052) on the clustering of clinical features in von Hippel-Lindau syndrome (HLS), Dr Filling-Katz and Dr Choyke, Dr Maher, and Professor Langenbeck (June 15, pp 1477-79) raise several interesting issues. Maher reports a heterozygote prevalence of 1 in 53 000 for HLS in East Anglia.This figure is in good agreement with our prevalence of 1 in 38 951, indicating that the Freiburg kindreds do not represent an epidemiological cluster of HLS. Apparently, there are occasional families presenting with both renal cell carcinoma and phaeochromocytoma. However, the great majority of recorded HLS kindreds show only one of these manifestations. An especially striking example is reported by Glenn et al.2 They observed an HLS family with 47 affected members, 57% of whom had phaeochromocytomas. Renal and pancreatic lesions were not detectable in any of these patients. In the same study most families presenting with renal and pancreatic features did not exhibit
phaeochromocytomas. Maher points out that
Department of Pathology, University of Zurich Zurich, Switzerland 1. Maher
Germany
HARTMUT P. H. NEUMANN
OTMAR D. WIESTLER
ER, Iselius L, Yates JRW, et al. Von Hippel-Lindau disease: a genetic study.
J Med Genet (m press). GM, Daniel LN, Choyke P, et al. von Hippel-Lindau disease: distinct phenotypes suggest more than one mutant allele at the VHL locus. Hum Genet (in press). 3. Neumann HPH. Basic criteria for clinical diagnosis and genetic counselling in von Hippel-Lindau syndrome. J Vasc Dis 1987; 16: 220-26. 4. Neumann HPH, Eggert HR, Weigel MD, et al. Hemangioblastoma of the central nervous system: a 10-year study with special reference to von Hippel-Lindau syndrome. J Neurosurg 1989; 70: 24-30. 5. Neumann HPH, Dinkel E, Brambs H, et al. Pancreatic lesions in the von Hippel-Lindau syndrome. Gastroenterology (in press) 2. Glenn
Fetal
DMD
family.
Arrow indicates carrier at risk.
renal carcinomas are frequently not detectable before the age of 44 (mean age of diagnosis in his study) and may, therefore, be missed in younger HLS patients. This cannot account for our results, since 12/28 of our patients from kindreds without renal lesions have passed the age of 44. Maher expresses concern that our focus on clustered manifestation of HLS may distract the attention for other features of the syndrome. All patients and potential gene carriers underwent a broad screening examination. 3,4 Filling-Katz and Choyke describe families with additional islet cell tumours and pancreatic adenocarcinoma as yet another example of a cluster of HLS features. However, it remains to be determined if these manifestations, which are most unusual in our HLS patients,s really represent manifestations of the syndrome. Several genetic models could account for the distinct patterns of features in HLS. We agree that, besides a complex genetic locus with several transcriptional units or cooperating genes on different chromosomes, allelic heterogeneity of the HLS gene on chromosome 3p is another potential genetic mechanism. Only a detailed molecular analysis in representative kindreds of the still elusive HLS gene will allow us to identify the genetic basis of these heterogeneous features. Department of Medicine, Albert Ludwig’s University, 7800 Freiburg im Breisgau,
Fig 1-Pedigree of Dutch
recombination distal to the DMD gene betwee H and F. However, the woman seeking advice had inherited an X-chromosome with a
potentially intragenic recombination between the proximal intragenic probe L and the first informative distal probe F. Although she had normal creatine kinase values, her carrier risk was high, possibly 20%. First trimester prenatal diagnosis on chorionic villi of her male pregnancy (III-2) showed the at-risk haplotype and the parents opted for termination. Muscle tissue of the fetus was studied immunohistochemically for dystrophin expression. We used monoclonal antibodies against the 30 kDa segment of dystrophin2 (aminoacids 1181-1388),3a59 kDa segment (aminoacids 17502248),4or the carboxy-terminal 17 aminoacids. All three showed clear staining of dystrophin in parts of the plasma membrane and at the distal ends of the myotubes5 (fig 2), an immunohistochemical pattern indistinguishable from that routinely found in normal fetuses. In affected fetuses, dystrophin cannot be detected with antibodies reactive with the distal part of the protein and sometimes not with
central antibodies either.6 Thus, this fetus was not affected and the recombination event in 11-2 has not included the DMD mutation. Her carrier risk was reduced to the population risk. Bieber et aF demonstrated by negative western blot testing that a 23-week-old DMD at-risk fetus was affected, confirming the DNA diagnosis and raising the mother’s carrier risk to 100%. We show here that even in younger fetuses sensitive dystrophin immunohistology permits genetic counselling. This will be especially valuable in DMD families where no mutation is found and the DNA markers are uninformative or show intragenic recombinations.
dystrophin to diagnose carrier status
SIR,-In a Dutch family with Duchenne muscular dystrophy (DMD) we have excluded carriership for a woman at risk (11-2, fig 1) by immunohistochemical analysis of her 12-week-old fetus after abortion for increased DMD risk. In this family, DNA analysis was uninformative. Two relatives (11-3,11-5) were affected with DMD. cDNA and polymerase chain reaction studies failed to show a mutation in the DMD gene, so the at-risk chromosome had to be identified by haplotype analysis.The informative markers used (H, F, L, C, and T) are identified in fig 1. Two non-carrier daughters were identified; 11-4 had the unaffected haplotype and 11-6 had a
Fig 2-lmmunohistological studies of fetal muscle. Tissue fixed and stained" and sections
were
carboxy-terminal monoclonal antibody.
was
immunostained with
