| BackgroundPart I NeurofibromatosisNeurofibromatosis, also known as Von Recklinghausen’s disease, is characterized by dysplasia of nervous system, skeletal system and skin.Clinical classificationClinically, neurofibromatosis can be classified into seven different types, including Neurofibromatosis type-1(NF1), NF2, NF3, NF4, NF5, NF6and NF7. NF1is the most common, counting for85%of all the cases, characterized by multiple cafe-au-lait, skinfold freckling, Lisch nodules, and neurofibromas, and involvement in central nervous system (CNS) is not common. NF2, mainly involving in CNS, is bilateral acoustic neuroma, presenting few cafe-au-lait and neurofibromas, and no Lisch nodules. NF3combines the features of both NF1and NF2. The types from NF4to NF7are very rare.ManifestationsCutaneous manifestations Skin pigment spots:The Cafe-au-lait macule (CALM) is one of the seven cardinal diagnostic criteria of NF1CALMs are discrete, well circumscribed, round or oval, uniformly pigmented patches. Despite the analogy of the color to coffee with milk, the pigmentation varies from light to dark brown. To fulfill this requirement, patients need six or more macules>5mm (prepuberty) or>15mm (postpuberty) in diameter. Less than1%of children under5years of age without NF1have more than two; when multiple macules are present, this is highly suggestive of NF1Another study found that the majority of patients with6or more CALMs will eventually meet diagnostic criteria for NF1, typically by age6years, and this likelihood increases with increasing number and typical morphologic appearance of CALMs. The prevalence of CALMs in the general population has varied between3%and36%-depending on the population studied-but the presence of multiple CALMs in an otherwise normal population is generally1%.Skin fold freckling, also known as Crowe’s sign, is the most specific of the cardinal criteria for NF1. It is considered nearly pathognomonic. But another study recently proposed that freckling may be seen in some patients with piebaldism and does not necessarily represent coexistence of neurofibromatosis type1. It is second only to CALMs in terms of age-related frequency and generally occurs between3and5years of age in either the axillae and/or groin. Other sites include under the neck and breasts, around the lips, and even the trunk in adults; however, none of these fulfill the NIH diagnostic criterion. Their size varies from1to3mm, distinguishing them from CALMs. Generalized hyperpigmentation also is one of skin pigmentary lesions of NF1.Cutaneous neurofibromas. The neurofibroma is considered as another hallmark sign of NF1. neurofibromas are the most common manifestations of neurofibromatosis type-1. They occasionally cause pain or progressive loss of function due to nerve compression. Neurofibromas can occur anywhere on the body and widely varies in their shape and size. Cutaneous neurofibromas usually do not become apparent until puberty and may continue to increase in size and number throughout adulthood. Outside of puberty, pregnancy is the other major time associated with increased growth. There are three types including skin one, hypodermal one and plexiform one. The latter occurs at birth and is important for diagnosis.Other cutaneous findings include juvenile xanthogranuloma, Melanoma, triad of juvenile xanthogranuloma (JXG), NF1, and juvenile myelomonocytic leukemia (JMML).Noncutaneous manifestationsOral lesions include oral tumor, at the site of the palate, buccal mucosa, tongue and the lips; or show acroglossia.NeuropathyNeuropathy, possibly localized or diffuse, central or peripheral, includes acoustic neuroma, optic glioma, hydrocephaly, heterotopic brain, glial nodule, neural tube closure, bulging meningomyelocele, brain and spinal cord tumors, schwannoma, ependymoma and astrocytoma and meningioma. Peripheral nerve lesions can appear abnormal sensation, nerve root pain or brachial plexus paralysis, epilepsy, mental retardation, ataxia, etc.Ocular disordersOcular disorders, such as lisch nodules, optic gliomas and anterior segment defects, are typical with clinical presentation of NF1. Anophthalmia is a rare eye malformation in NF1. Ophthalmic features of NF2are also prominent and include reduced visual acuity and cataract. The two most common problems are lisch nodules, optic gliomas.Lisch nodules are small, dome-shaped hyperpigmented macules of the iris that cause no impairment of vision. They are a common finding (by6years of age,15%to20%of children have them;95%of adults have them) and are included as one of the cardinal NIH diagnostic criteria.Bony abnormalitiesBony abnormalities in NF1are variable and include scoliosis, sphenoid wing or long bone dysplasia, and, more recently, osteopenia/osteoporosis. Patients with NF1display significantly lower25-(OH)-cholecalciferol serum levels and decreased bone mineral density(BMD). Histomorphometric analysis not only reveal a reduced trabecular bone volume in biopsies from NF1patients, but also a significantly increased osteoid volume and increased numbers of osteoblasts and osteoclasts. Moreover, qBEI analysis revealed a significant decrease of the calcium content in biopsies from NF1patients.5. Endocrine abnormalities lead to acromegaly, myxedema, precocious puberty or delay, Addison’s disease, hyperparathyroidism, medullary thyroid carcinoma and chromaffin cells carcinoma.MalignancyNeurofibrosarcoma, also known as malignant schwannoma, can occur on patients with neuroflbromatosis, but it is not common. In the skin, the tumor is named as malignant peripheral nerve sheath tumor(MPNST)and rarer.HostopathologicallyCafe-Au-Lait Maecula Increased pigment is seen in keratinocytes and melanocytes of epidermis. Giant global melanin granules may be seen in melanocytes and basal cells with diffused distribution. The granules are about5μM in diameter, and Dopa reaction shows increased density and activities of melanocytes.Neurofibroma No envelope exists on the tumors, which may spread to the hypodermal fatty tissues and have a clear bound, composed of nerve garment and Schwann cells. Within the tumor, hyperplasia of neural axon and richness of small blood vessels is showed. Schwann cells are in long and thin spindle shape or slightly bent into waveform with unclear boundary. Plasma of the cells is stained in color of light eosin.They have an obviously different length on both ends, are arranged in waveform or turbine with few fibroblasts in it. Special staining frequently shows mast cells and no elastic fiber. Plexiform neurofibromatosis involves in deep large nerve, in which irregular nerve tracts are seen. The early-stage local nerve envelope appears myxoid, more interstitial substances and have an invasive, and makes medullated and non-medullated neuraxon divide. Nerve sheath cells and collagen fiber hyperplasia forms bending cords, surrounded by myxoid amorphous stroma. Morphologically, the normal nerve can not be distinguished at advanced stage due to enlargement in size of tumors.Diagnostic criteriaBased on the1988National Institutes of Health Consensus Development Conference on Neurofibromatosis, the diagnosis of NF1is made in an individual with any2of the following clinical features:(1)6or more cafe-au-lait spots,(2) axillary or groin freckling,(3)2or more Lisch nodules,(4)2or more neurofibromas,(5) optic pathway gliomas (OPGs),(6) bone dysplasia, and (7) a first-degree family relative with NF1The diagnosis of NF2is made in an individual with any1of the following clinical features:(1) the tests of CT and MRI show tumor involving in the bilateral vestibular nerves;(2) a first-degree family relative with NF2and at least one side eighth of cranial nerves is involved, or2or more tumors as following:neurofibroma, meningioma, glioma and schwannomManagement Up to now, there is no perfect therapy for treating neurofibromatosis. Surgical intervention is the only effective means of treatment for progressive pain, disfigurement, functional compromise, and malignancy. Complete surgical resection is the standard treatment for malignant peripheral nerve sheath tumors. Cafe-au-lait spots can be treated by laser therapies, including pulse-dye laser, YAG laser and ruby laser. Kim, H. J. et al. reported a successful treatment of multiple cutaneous neuroflbromas using a combination of shave excision and laser photothermocoagulation with a1,444-nm Neodymium-Doped Yttrium Aluminum Garnet (Nd:YAG) laser, carefully checking lesions should be taken if of onset of epilepsy. If necessary, neurosurgery may be considered to resect the lesions which is possibly recurrent.Part Ⅱ Neurofibromatosis type1Neurofibromatosis type1(NF1; OMIM#162200) is a common neurocutaneous disorder that is characterized by multiple cafe-au-lait, skinfold freckling, Lisch nodules, and neurofibromas. Mutations in the NF1gene, which encodes the neurofibromin protein, have been identified as the pathogenic gene of NF1. The incidence has been estimated to be1in3500individuals. The less frequent manifestations include malignancies, plexiform neurofibromas, optic glioma, learning difficulties, juvenile xanthogranuloma and skeletal abnormalities. About50%of all malignant peripheral nerve sheath tumors (MPNSTs) arise as neurofibromatosis type1associated lesions.Cafe-au-lait (CAL) spots occur in more than90%of NF1cases, and manifested as well-circumscribed, light-brown, homogenous patches that range from1to2mm to greater than20cm in diameter, with the majority being under10cm. A giant cafe-au-lait macule with irregular borders, involving the lower part of the back, right flank and both buttocks and thighs were reported recently, which is rather unusual. Neurofibromas, another one of the hallmark signs of NF1, can occur anywhere on the body and there is a wide variation in their shape and size. Generally, the’cutaneous’or’dermal’tumors are dome-shaped, soft, fleshy, and skin colored to slightly hyperpigmented, and the’subcutaneous’ tumors are of the firm, nodular variety. Neurofibromas, beside skin, also appear in the gynecologic tract, including the cervix, resulting in cervical stenosis and lower abdominal pain.NF1gene, mapped on chromosome17ql1.2, spans approximately280kb of genomic DNA, contains60exons and encodes neurofibromin which is composed of2818amino acid residues. Neurofibromin that functions as a GTPase activating protein (GAP) for Ras is widely expressed in many tissues. To date, more than1,000different NF1mutations have been found and listed in the Human Gene Mutation Database. The majority of these NF1mutations lead to truncated protein products.ObjectivesTo identify the mutations of NF1gene and the correlation of genotype and phenotype in human samples on the basis of research results of model organisms for further illuminating the biological function of NF1gene, expand the data of NF1gene mutation and proivide a foundation of genetic consultation, prenatal diagnosis, genetic diagnosis and the future genetic treatment in neurofibromatosis type1.Materials and methodsThe NF1gene of two families with neurofibromatosis type1and three sporadic cases and100unrelated controls were amplified by polymerase chain reaction(PCR) and then were sequenced. The genes of subjects of the study were analysed by software BioEdit. The correlation study between the genotypes and phenotypes in these patients was exerted by comparing the data of clinic and mutations.resultsClinical findingsPatient1, a3.5-year-old girl, was the proband in family1. She presented with cafe-au-lait spots involving trunk, extremities and buttocks. Her father, aunt, grandmother have more typical and severe manifestation of many neurofibromas and cafe-au-lait spots over their bodies. In the family, all patients had cafe-au-lait spots at birth, and the neurofibromas appeared in5or6years old, and rapidly developed at teenage age. The other three patients [the proband in family2, and the two sporadic patients] also had the typical clinical features of neurofibromatosis. In family2, all patients had cafe-au-lait spots at birth, and the neurofibromas developed in about3years old, and gradually developed at teenage age. Patient3was a46year-old male, sporadic case, who had onset at the age of20s. However, the neurofibromas developed very swiftly and the size of the tumors was very giant partly. On histological examination of skin biopsies taken from one patient, the typical features of neurofibromas and spindle cell proliferations were seen, with a mucinous background and many mast cells.Experimental findingsThe sequencing of PCR products showed three novel mutations in NF1gene, c.601T>A in exon4, c.871G>T in exon6, c.1448A>G in exon10, respectively leading to amino acid changes, F201I, E291X, D485G. No any change was found in unaffected individuals in these families and unrelated controls.Conclusions(1) These mutations c.601T>A, c.871G>T, c.1448A>G of NF1gene may be pathogenic causes for neurofibromatosis type1.(2) Our data indicates that the mutations in exon4,6,10of NF1gene possibly change the structure and function of NF1protein and ultimately affect their physiological function. |
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