The Study Of Clinical And Pathological In Children With Alport Syndrome In Southwest Of China

PART ONE: USE OF PARAFFIN-EMBEDDED RENAL SECTIONFOR IMMUNOFLUORESCENCE STAINING IN ALPORTSYNDROMEObjective To establish the diagnostic technique of Alportsyndrome(AS) by immunofluorescence staining of type Ⅳcollagen α chainson paraffin-embedded renal sections．Methods4patients with X-linked dominant form of AS（3males and1females） were retrospectively analyzed，who were admitted betweenApril，2012to January，2013． AS was diagnosed according to clinicalsymptoms，family history，pathology，immunofluorescence staining of typeIV collagen chains on renal. Normal portions of renal tissue from4patientswith renal duplication were used as controls．Type Ⅳ collagen α5chainswere stained by indirect immunofluorescence staining method onparaffin-embedded renal sections． Three antigen retrieval methods,including microwave treatment，pepsin digestion and microwave treatment plus pepsin digestion were first used in the controls, to find the best antigenretrieval method for type Ⅳ collagen α5chains．The results were comparedwith immunofluorescence staining on fresh frozen sections from the sameset of cases to evaluate the methodology.Results The type Ⅳ collagen α5chains immunofluorescence stainwere negative on formaldehyde-fixed tissue and paraffin-embedded sectionspreceded by microwave treatment or pepsin digestion epitope retrievalalone, but the paraffin-embedded renal sections preceded by microwavetreatment after pepsin digest10minutes, type Ⅳ collagen α5chains showedcontinuous linear pattern along glomerular basement membrane（GBM） onsections from the controls, negative for X-linked dominant male AS patients,intermittent linear pattern for X-linked dominant female AS patient. Thesame results as immunofluorescence on frozen tissue.Conclusions XLAS can be diagnosed by immunofluorescencestaining of type Ⅳ collagen α5chains on paraffin-embedded renalsections.Moreover，it can make those Previously suspect XLAS diagnosis byclinicopathology or renal biopsy electron microscope to be definite，which isa useful technique for diagnosis of XLAS. PART TWO:CLINICAL AND PATHOLOGICAL FEATURES OFCHILDREN WITH ALPORT SYNDROME IN SOUTHWEST OFCHINAObjective To analyze the clinical and pathological features of childrenwith Alport syndrome in southwest of China.Methods20patients with AS of southwest China were retrospectivelyreviewed，the clinical data was collected. The distribution of type IVcollagen α chain in renal and skin tissue was detected by indirectimmunofluorescence assay. Clinical and pathological features of childrenwith AS were compared in different clinical phenotypes as well.Results Most of the children showed microscopic hematuria associatedwith proteinuria as Initial symptoms. The level of protein in24h urinary washigher in patients with isolated proteinuria than that in those with hematuriaand proteinuria (P<0.05). The proportion of uremic patients in the family ofthe children with Proteinuria is (6/16) more than that in the family ofchildren with hematuria (1/4), but no significant difference. The findings bylight microscope mostly revealed mild to moderate mesangial proliferativeglomerulonephritis (MsPGN)(15/20). There was no significant differencein Children AS with variable clinical phenotypes on pathological typedistribution (P>0.05). About60%of the AS existed foam cells of kidneyinterstitium, of which the urine protein level was significantly higher than those with non-foam cells (P<0.05). Based on the different AS genotypes’immunofluorescence characters of type Ⅳ collagen α chains in basemembrane,15patients (75%) were diagnosed as X-linked dominantinherited AS (XLAS), and5patients were still not yet determined theinherited types according to the deletion of type IV collagen α5chains. Twochildren with AS were also detected of collagen type IV α5chain inEpithelial basement membrane (EBM), the results of immunofluorescencestaining were negative in male and discontinuous or mosaic pattern infemales(whose mother also exhibited discontinuous or mosaic pattern in theimmunofluorescent staining of the epidermal basementmembrane). Thetwo patients were diagnosed as XLAS, the same result as detecting type Ⅳcollagen α5chains in GBM.Conclusions In Southwest China, the clinical manifestation wasvariegated in children with Alport syndrome, and microscopic hematuriaassociated with proteinuria appeared in most patients. The main inheritedtype was XLAS (75%). Though there was no significant difference inChildren AS with variable clinical phenotypes on pathological typedistribution, the higher the level of proteinuria, the more the patient hadsevere change in renal tubular interstitial. The diagnosis of XLAS by skinbiopsy showed the same results as detecting by renal, if necessary, it canreplace renal biopsy under certain circumstance. Moreover, it plays an important role in screening the carriers and making genetic counseling.