الفهرس 
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Abstract
Nephrotic syndrome (NS) is a common childhood kidney disease caused by impaired glomerular function, characterized by protein leakage from the blood to the urine through the glomeruli, resulting in proteinuria, hypoalbuminemia, hypercholesterolemia and generalized oedema. NS is descriptively classified upon the patients response to steroid treatment as steroid sensitive NS (SSNS) or steroid resistant NS (SRNS).
In addition, the steroid-resistant nephrotic syndrome is caused by abnormal receptor genes as a drug reaction and develops into total renal failure due to the absence of an early-stage gene therapy protocol.
1) The MDR-1 gene encodes multidrug transporter P glycoprotein (P-gp) which functions as a transmembrane efflux pump; therefore it is important in the absorption, tissue targeting, and elimination of different drugs, It is a multi-genetic form G2677T / A and C3435T is significantly higher in children with steroid-resistant nephrotic syndrome compared to healthy children and C2677T / A is also significantly higher in patients with steroid-resistant nephrotic syndrome than patients with steroid-sensitive nephrotic syndrome. 2) Interleukin 6 (IL-6) and genetic variants have been studied in various disorders with recurrent infections and immunological changes such as gastrointestinal disorders, lupus erythromatosus, and leishmaniasis. Therefore, IL6 G174C is more prevalent in patients with steroid resistance nephrotic syndrome, and therefore can be responsible for the development as well as for steroid resistance.
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3) TNF-α has a strong immune function Previous studies have shown increased TNF-α synthesis and gene expression in patients with idiopathic nephrotic syndrome and focal segmental glomerulo sclerosis (FSGS).
This work aims to study: MDR1 gene polymorphism and TNFα and IL6 gene polymorphism in steroid resistant nephrotic syndrome in Egyptian children.
All subjects included in the study will be recruited from Mansoura University Children’s Hospital. The Ethics Committee of Faculty of science, Port Said University approved our study protocol, and children’s parents assigned written informed consent.
The study will be concluded on two hundred (250) subjects divided into three groups.
group 1:100steroid resistant nephrotic syndrome (SRNS) patients with age (range:6-18 years).
group 2:50 steroid sensitive nephrotic syndrome (SSNS) patients act as control with age (range: 6-18 years).
group 3:100 healthy children act as healthy control with age (range: 6-18 years).
Blood and Urine sampling:
Blood samples were drawn from all subjects both patients and controls after an overnight fast and divided into 2portions:
2 ml of whole blood was collected into tubes containing EDTA, for genomic DNA extraction; polymerase chain reaction (PCR) and Restriction fragment length polymorphism (RFLP).
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3 ml of blood was added to polypropylene tubes with stopper, left to clot for 20 minutes at 37 °c centrifuged at 3000 xg for 10 minutes. The resulting serum was separated from the sample and stored at−20 °C until analysis.
The urine samples were collected from each participant
All children were subjected to the following:
1) Complete history and physical examination.
2) Laboratory investigations.
a) Routin laboratory investigation: serum creatinine, cholesterol, albumin, blood urea nitrogen and urine protein. b) Serum interleukin-6 (IL-6) and tumor necrosis factor- alpha (TNF-α) by ELISA technique.
c) The multi drug resistance gene (MDR1) gene polymorphism by polymerase chain reaction (PCR) - restriction fragment length polymorphism (RFLP). d) Interleukin-6 (IL-6) and tumor necrosis factor- alpha (TNF-α) gene polymorphism by PCR-RFLP.
The obtained results summarized the following:
1- Creatinine:
Creatinine showed significantly higher concentrations in NS cases when compared to control group (P<0.001). On other hand creatinine showed no significant differences were found in SRNS when compared to SSNS subgroups.
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2- Cholesterol:
Cholesterol showed significantly higher concentrations in NS cases when compared to control group (P<0.001). Also cholesterol showed significantly higher concentrations in SRNS when compared to SSNS subgroups (P<0.001).
3- Albumin:
Albumin showed significantly lower concentrations in NS cases when compared to control group (P<0.001). Also albumin showed significantly lower concentrations in SRNS when compared to SSNS subgroups (P=0.045).
4- Blood urea nitrogen:
Blood urea nitrogen showed significantly higher concentrations in NS cases when compared to control group (P<0.001). On other hand blood urea nitrogen showed no significant differences were found in SRNS when compared to SSNS subgroups.
5- Urine protein concentration:
Urine protein concentration showed significantly higher concentrations in NS cases when compared to control group (P<0.001). But, there is no significant differences were found in urine protein concentration between SRNS and SSNS subgroups.
6- Serum IL6 and TNF α concentration:
IL6 and TNF α showed significantly higher concentrations in NS when compared to healthy control group (P<0.001), as well as in SRNS when compared to SSNS subgroups (P<0.001).
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-The multi drug resistance gene (MDR1) gene polymorphism byPCR-RFLP:
Our results suggest that, MDR C3435T TT, CT+TT genotypes, T allele(P = 0.028, P = 0.015 and P = 0.001 respectively) and MDR G2677A/T GT+GA, TT+AA, GT+GA+ TT+AA genotypes, T+A allele (P <0.001 for all) showed significant differences between NS and healthy control groups, MDR C1236T genotypes and alleles did not differ significantly between NS and control groups.
MDR G2677A/T GT+GA, TT+AA, GT+GA+TT+AA genotypes, T+A allele showed significant differences between SDNS and SRNS groups, with risk to develop SRNS within NS patients, even after adjustment with age and gender (P = 0.001, P = 0.003, P <0.001 and P = 0.002 respectively). MDR C3435T and C1236T genotypes and alleles did not differ significantly between SSNS and SRNS groups. -Interleukin-6 (IL-6) and tumor necrosis factor- alpha (TNF-α) gene polymorphism by PCR-RFLP:
Our results revealed that, IL6 GC, CC, GC+CC genotypes, C allele (P< 0.001, = 0.008, < 0.001 &< 0.001 respectively) and TNF GA, AA, GA+AA genotypes, A allele (P=0.003, P=0.027, P=0.001 & P< 0.001 respectively) showed significant association with risk of NS development within healthy control subjects even after adjustment with age and gender.
Also IL6 CC, GC+CC genotypes, C allele showed significant association with risk of SRNS development within NS subjects even after adjustment with age and gender (P = 0.022, = 0.045 & = 0.022 respectively).
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On other hand, TNF genotypes and alleles did not differ significantly between SSNS and SRNS groups.