OBJECTIVETo screen the 5' regulatory region of the aldose reductase (AR) gene for genetic variabilities causing changes in protein expression and affecting the promoter function.

METHODSThe screenings were carried out by polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP). All SSCP variants were submitted for DNA sequencing and inserted into the plasmid chloromycetin acetyl transferase (CAT) enhancer vector. The constructs were used to transfect Hela cells, and CAT assays were performed to assess promoter activity. Gel mobility shift and footprinting assays were also performed to determine the interaction between the DNA and nuclear proteins.

RESULTSTwo polymorphisms, C (-106) T and C (-12) G, were identified in the regulatory region in 123 Chinese control subjects and 145 patients with type 2 diabetes mellitus. The frequencies of genotypes WT/WT, WT/C (-12) G and WT/C (-106) T were not significantly different between the subjects and patients. In the patients with and without retinopathy, frequencies of WT/C (-106) T were 31.5% and 17.5% (P < 0.05) respectively, and the frequencies of WT/C (-12) G were 10.5% and 2.5% (P > 0.05) respectively. The total frequency of WT/C (-12) G and WT/C (-106) T in patients with retinopathy was 41.8%, significantly higher than that (20.0%) in patients without retinopathy (P < 0.025). The relative transcription activities of the wild-type, the C (-12) G and the C (-106) T were 15.7%, 31.0% and 32.2%, respectively. The results of DNA-protein interaction assays showed that these variations did not change the binding site of DNA with trans-acting factors.

CONCLUSIONThe polymorphisms C (-12) G and C (-106) T strongly associated with diabetic retinopathy in the Chinese population have been identified in the regulatory region of the aldose reductase gene.

5' Flanking Region ; genetics ; Adult ; Aldehyde Reductase ; genetics ; metabolism ; Binding Sites ; genetics ; China ; Chloramphenicol O-Acetyltransferase ; genetics ; metabolism ; DNA ; chemistry ; genetics ; DNA Footprinting ; Diabetes Mellitus, Type 2 ; enzymology ; genetics ; Electrophoretic Mobility Shift Assay ; Female ; HeLa Cells ; Humans ; Male ; Middle Aged ; Mutation ; Polymerase Chain Reaction ; Polymorphism, Genetic ; Polymorphism, Single-Stranded Conformational ; Recombinant Fusion Proteins ; genetics ; metabolism ; Regulatory Sequences, Nucleic Acid ; genetics ; Sequence Analysis, DNA ; Transcription, Genetic

Objective:To explore the advantages, disadvantages, and precautions of clinically applying four types of non-trunk vessels as recipient vessels in the free flap transplantation for repairing peri-knee wounds.Methods:A retrospective analysis of the clinical data was conducted of 23 patients (12 males and 11 females) with peri-knee skin and soft tissue defects who underwent free anterolateral thigh flaps or free latissimus dorsi flaps. The patients were admitted to the Department of Traumatic Orthopedics, Wuxi 9th People’s Hospital, from January 2015 to December 2019. The patients were aged 20-72 years (mean 41.9). The wound size with vital tissue exposure was 18.0 cm×5.0 cm-42.0 cm×9.0 cm. Preoperative color Doppler and computerized tomography angiography techniques were used to assist in positioning and to measure the recipient vessels (including the descending genicular vessel, descending branch of the lateral femoral circumflex vessel, the medial sural vessel, and the medial inferior genicular vessel) and blood vessels in the donor area (including descending branches of lateral femoral circumflex vessels and thoracic dorsal vessels). The caliber was measured and verified by a soft ruler with a scale intraoperative. The measured values of the caliber were recorded in the preoperative and intraoperative. The color and texture of the two flaps and the healing of the donor sites were observed postoperatively. The conformance ratio of preoperative and intraoperative measured values of vessels in the donor and recipient areas (except for the only case of the medial inferior genicular vessel) was compared. SPSS 26.0 software was used for data analysis. The measurement data were showed as Mean±SD, and the comparison results were analyzed by paired t-test. P<0.05 was considered statistically significant. Results:A total of 23 patients with skin and soft tissue defects around the knee were enrolled. The size of the tissue flap was 18.0 cm×5.0 cm-46.0 cm×9.0 cm. Twelve of 14 anterolateral thigh flaps anastomosed to the descending genicular vessel survived. The remaining two cases had a length of 6 cm and 4 cm necrosis at the distal flap, which was finally given skin-grafting and healed. One of the 12 survived flaps failed in limb salvage in Phase Ⅰ and was thus given flap reconstruction in Phase Ⅱ. Another case was given amputation due to serious infection of limbs, with incomplete ends of the survived flaps rotated and covered. In four cases anastomosed to the descending branch of the lateral circumflex femoral vessel, three anterolateral thigh flaps survived completely, and one distal latissimus dorsi flap had a length of 12 cm necrosis which was given debridement and Ilizarov bone transport for heal. Two anterolateral thigh flaps and two latissimus dorsi flaps anastomosed to the medial sural vessel survived completely, of which one anterolateral thigh flap had vein crisis which was later solved. The anterolateral thigh flap of 1 case anastomosed to medial inferior genicular vessels completely survived. The postoperative follow-up lasted 3-30 months with an average of 13.6 months. All the flaps have good color and textures with good incision heal at the donor site in Phase Ⅰ. There was no statistically significant difference in the preoperative and intraoperative measurement values of blood vessel caliber in the donor and recipient areas ( P>0.05). Conclusions:Four non-trunk peri-knee blood vessels can serve as recipient vessels of the free tissue flaps, and proper selection of the vessels can effectively improve the survival rate of the tissue flaps. The descending genicular vessel can serve as the recipient vessel for a priority, as with a superficial position, fixed dissection, simple positioning, and convenient intraoperative dissection.

Objective:To explore the advantages, disadvantages, and precautions of clinically applying four types of non-trunk vessels as recipient vessels in the free flap transplantation for repairing peri-knee wounds.Methods:A retrospective analysis of the clinical data was conducted of 23 patients (12 males and 11 females) with peri-knee skin and soft tissue defects who underwent free anterolateral thigh flaps or free latissimus dorsi flaps. The patients were admitted to the Department of Traumatic Orthopedics, Wuxi 9th People’s Hospital, from January 2015 to December 2019. The patients were aged 20-72 years (mean 41.9). The wound size with vital tissue exposure was 18.0 cm×5.0 cm-42.0 cm×9.0 cm. Preoperative color Doppler and computerized tomography angiography techniques were used to assist in positioning and to measure the recipient vessels (including the descending genicular vessel, descending branch of the lateral femoral circumflex vessel, the medial sural vessel, and the medial inferior genicular vessel) and blood vessels in the donor area (including descending branches of lateral femoral circumflex vessels and thoracic dorsal vessels). The caliber was measured and verified by a soft ruler with a scale intraoperative. The measured values of the caliber were recorded in the preoperative and intraoperative. The color and texture of the two flaps and the healing of the donor sites were observed postoperatively. The conformance ratio of preoperative and intraoperative measured values of vessels in the donor and recipient areas (except for the only case of the medial inferior genicular vessel) was compared. SPSS 26.0 software was used for data analysis. The measurement data were showed as Mean±SD, and the comparison results were analyzed by paired t-test. P<0.05 was considered statistically significant. Results:A total of 23 patients with skin and soft tissue defects around the knee were enrolled. The size of the tissue flap was 18.0 cm×5.0 cm-46.0 cm×9.0 cm. Twelve of 14 anterolateral thigh flaps anastomosed to the descending genicular vessel survived. The remaining two cases had a length of 6 cm and 4 cm necrosis at the distal flap, which was finally given skin-grafting and healed. One of the 12 survived flaps failed in limb salvage in Phase Ⅰ and was thus given flap reconstruction in Phase Ⅱ. Another case was given amputation due to serious infection of limbs, with incomplete ends of the survived flaps rotated and covered. In four cases anastomosed to the descending branch of the lateral circumflex femoral vessel, three anterolateral thigh flaps survived completely, and one distal latissimus dorsi flap had a length of 12 cm necrosis which was given debridement and Ilizarov bone transport for heal. Two anterolateral thigh flaps and two latissimus dorsi flaps anastomosed to the medial sural vessel survived completely, of which one anterolateral thigh flap had vein crisis which was later solved. The anterolateral thigh flap of 1 case anastomosed to medial inferior genicular vessels completely survived. The postoperative follow-up lasted 3-30 months with an average of 13.6 months. All the flaps have good color and textures with good incision heal at the donor site in Phase Ⅰ. There was no statistically significant difference in the preoperative and intraoperative measurement values of blood vessel caliber in the donor and recipient areas ( P>0.05). Conclusions:Four non-trunk peri-knee blood vessels can serve as recipient vessels of the free tissue flaps, and proper selection of the vessels can effectively improve the survival rate of the tissue flaps. The descending genicular vessel can serve as the recipient vessel for a priority, as with a superficial position, fixed dissection, simple positioning, and convenient intraoperative dissection.