Published ahead of print on April 12, 2007, doi:10.1164/rccm.200610-1408CR
© 2007 American Thoracic Society doi: 10.1164/rccm.200610-1408CR
Pulmonary Lymphangioleiomyomatosis in a Karyotypically Normal Man without Tuberous Sclerosis Complex1 Unit of Lung Physiopathology, and 2 Unit of Radiology, Azienda Ospedaliera S. Orsola-Malpighi and University of Bologna, Bologna, Italy; 3 Unit of Pathologic Anatomy, S. Maria Nuova Hospital, Reggio Emilia, Italy; 4 Department of Genetics, Biology, and Biochemistry, University of Turin, Turin, Italy; and 5 Institute of Thoracic Surgery, and 6 Unit of Pathologic Anatomy, F. Addarii Institute of Oncology, Azienda Ospedaliera S. Orsola-Malpighi and University of Bologna, Bologna, Italy Correspondence and requests for reprints should be addressed to Antonia D'Errico-Grigioni, M.D., Istituto di Oncologia F. Addarii, Viale Ercolani 4/2, 40138 Bologna, Italy. E-mail: derrico{at}aosp.bo.it
Rationale: The three previously reported cases of conclusively documented pulmonary lymphangioleiomyomatosis (LAM) in men were associated with definite or probable tuberous sclerosis complex (TSC). Objectives: To describe an unequivocal case of pulmonary LAM occurring in a man with no clinical or genotypic evidence of TSC. Methods: At high-resolution computed tomography, a 37-year-old phenotypically and karyotypically normal man with left pneumothorax and massive pulmonary collapse had widespread thin-walled cysts throughout both lungs. Histological diagnosis of LAM was performed on biopsy material, and immunohistochemically confirmed with the HMB-45 monoclonal antibody. Measurements and Main Results: Remarkably, the HMB-45–positive cells lining the cysts also showed strong reactivity for estrogen and progesterone receptor proteins. TSC was clinically excluded, and TSC1 and TSC2 germline mutations were not detected at DNA analysis. Conclusions: This article indicates that occurrence of LAM may be possible in a chromosomally normal man unaffected by TSC. On diagnostic grounds, the possibility of LAM should be borne in mind when diffuse cystic lung disease occurs in a man, even in the absence of signs of TSC.
Key Words: smooth muscle cells • histopathology • sex distribution • rare diseases Pulmonary lymphangioleiomyomatosis (LAM) is a rare interstitial disease of uncertain etiology, which is almost exclusively confined to women and is frequently associated with tuberous sclerosis complex (TSC) (1–5). Histologically, LAM is characterized by proliferation of peculiar smooth muscle cells (LAM cells) that show reactivity to the human melanoma black (HMB)–45 monoclonal antibody (6) in the bronchioles, pulmonary veins, and lymphatic vessels. A systematic review of the literature found reports of only nine men bearing a possible, but unproven, diagnosis of pulmonary LAM between 1939 and 1997 (7). A current search of PubMed and Embase identified three subsequent reports of pulmonary LAM in men, all with either definite (8, 9) or probable (10) TSC. Here, we describe an unequivocal case of pulmonary LAM occurring in a man with no evidence at DNA analysis of TSC1 or TSC2 gene mutations.
In December 2003, a 37-year-old phenotypically normal man was urgently admitted to our hospital due to left pneumothorax and massive pulmonary collapse. The patient, who was a company manager and father of a normal child, denied having ever smoked and reported no personal history of serious illness or hospitalization, seizures, mechanical ventilation, or prematurity. Family anamnesis did not disclose possible cases of LAM, TSC, emphysema, pneumothorax, renal cancer, seizures, or mental retardation. Thoracic drainage was performed and the patient was discharged after 10 days. One month later, a high-resolution computed tomography (CT) scan revealed widespread thin-walled cysts (0.5–1.5 cm in diameter) throughout both lungs, showing the characteristic features of pulmonary LAM (Figure 1). Serological  1-antitrypsin values were normal. Clinical signs and laboratory tests for Sjögren's syndrome were negative. Brain and abdominal CT scans were negative for cerebral calcifications and angiomyolipomas; no subtle anomalies were apparent. Abdominal echography confirmed existing diagnoses of a 3-cm renal cyst (6 Hounsfield units at CT) and a 3-cm hepatic hemangioma in the VII segment (hyperechogenic with a clear border). Video-assisted thoracoscopic biopsies were taken at the lingula and apex of the left lung, and a pleural abrasion was performed. Histological examination and immunostaining (see Figure 2) confirmed a final diagnosis of LAM, with characteristic cytoplasmic reactivity to the HMB-45 monoclonal antibody in LAM cells lining the cysts. Remarkably, the same cells showed strong nuclear reactivity for estrogen and progesterone receptor proteins.
Regarding gender status, estrogen, progesterone, estrone, and testosterone values were all within the normal range. Physically, the patient had normal developmental features, including two descending testicles and male pattern frontal balding and hair distribution. Karyotyping demonstrated that the patient had a normal male complement of chromosomes: conventional GTG- and QF-banding (band resolution, 550) of metaphase chromosomes obtained from phytohemagglutinin (PHA)-stimulated blood lymphocyte cultures showed a normal 46,XY karyotype in 100 examined metaphases, thereby excluding the possibility of chimerisms. Clinical signs of TSC were systematically investigated. An expert dermatologist was unable to find signs of TSC, such as angiofibromas, subungual fibromas, hypopigmented macules (a Wood's lamp was used), or confetti lesions. An ophthalmologist was also unable to find signs of TSC. Mutation analysis on DNA from cyst wall tissue and peripheral blood lymphocytes was performed to search for TSC1 or TSC2 mutations. Of note, in February 2004, a decision was made based on the clinicians' judgment (and in concordance with the patient) to use hormonal manipulation therapy, a treatment option that has been tried in women using various pharmacological strategies (5) (we administered leuprorelin, a gonadotropin releasing hormone [GnRH]-antagonist analog, initially associated with letrozole, an aromatase inhibitor). At the time of writing (December 2006) the patient feels in normal health and appears to be clinically and functionally stable.
Histology and Immunohistochemistry Routine hematoxylin-and-eosin–stained sections of the lung biopsy were examined. Immunohistochemical stains for HMB-45 (1:30; BioGenex, San Ramon, CA), estrogen receptor (1:50, Clone 1D5; Dako, Glostrup, Denmark) and progesterone receptor (1:50, Clone PgR 636; Dako), smooth muscle actin (1:40, Clone HHF35; BioGenex), and desmin (1:200 Clone D33; Dako) were performed on a Dako Autostainer (Dako, Carpintera, CA) using a standard avidin–biotin complex (Dako, Glostrup, Denmark) method.
Screening of TSC1 and TSC2 Germline Mutations
Point mutation screening.
Screening for large deletions.
Immunohistochemistry LAM cells showed strong nuclear immunoreactivity for estrogen and progesterone receptor monoclonal antibodies, and cytoplasm staining for monoclonal antibodies against HMB-45 and smooth muscle actin (Figure 2).
DNA Analysis
To our knowledge, this is the first report of definite diagnosis of pulmonary LAM in a karyotypically normal man, apparently not affected by TSC (in addition to the negative expert clinical evaluation, TSC1 and TSC2 germline mutations were not detected). The present case suggests that TSC is not a necessary condition for onset of LAM in men. The only finding in our karyotypically normal man that could provide a pathogenetic clue was the presence of estrogen and progesterone receptor proteins in the LAM cells. This is a common finding in women affected by LAM, and a similar observation was recorded in one of the three cases previously reported in men (8) (the LAM cells of one of the two other men showed only mild positivity for progesterone receptors [10]). The negative results at DNA analysis require some comment. In the lung tissue, we did not identify any mutation in the TSC1 or TSC2 genes. For TSC1, loss of heterozygosity was excluded in exon 22, where a polymorphism was found in both blood and lung tissue (no polymorphism was detected in any exon of TSC2). Regarding sensitivity, reported detection rates range from 37 to 83%, with the best results deriving from DHPLC, long-range PCR, and quantitative PCR (12). Our current detection rate using MLPA, DHPLC, and sequencing is 75 to 80%. A plausible explanation for the minority (15–20%) of TSC cases who test negative is the existence of a mutation in regulatory regions of the genes, or in a deep intron not investigated by DHPLC. Because mechanical microdissection was used in the present case, it also cannot be ruled out that the presence of normal tissue might have masked mutations in the LAM cells (although we believe this is unlikely in view of the extreme scarcity of normal tissue in the specimens used). Knowledge that LAM may occur in a chromosomally normal man in the absence of TSC could be relevant for research into the etiology and pathogenesis of this rare disease. On diagnostic grounds, the possibility of LAM should be borne in mind when diffuse cystic lung disease occurs in a man, even in the absence of signs of TSC.
The authors would like to dedicate this article to the memory of Emanuele Grazia of the University of Bologna, who was one of the surgeons who worked on this case. His sudden death occurred while this manuscript was being prepared. Originally Published in Press as DOI: 10.1164/rccm.200610-1408CR on April 12, 2007 Conflict of Interest Statement: None of the authors has a financial relationship with a commercial entity that has an interest in the subject of this manuscript. Received in original form October 3, 2006; accepted in final form April 11, 2007
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