Purpose Ureteropelvic junction obstruction (UPJO) is the most common cause of congenital hydronephrosis. It was suggested an important role in smooth muscle cell (SMC) malfunction and apoptosis as primary anomaly in UPJO. SMC cytoskeleton in UPJO, however, has not been investigated. Vinculin-talin-integrin arrangement is a known systems of interaction between extracellular matrix and sarcolemma-associated cytoskeleton. Our purpose is to evaluate the cytoskeletal structure of SMC of infants affected by UPJO. Material and Methods Tissue specimens obtained from ten pyeloplasty were divided into 3 sections, renal pelvis above the obstruction, UPJ and ureter below the obstruction. For control group, eight unaffected UPJ of matched-aged infants were autopsied. Specimens, fixed in paraformaldeyde, were examinated using anti-β1-D, anti-β1-A, anti-α7-B and anti-α7-A integrins, anti-talin, anti-vinculin and anti-β-dystroglycan antibodies. Sections were observed using a confocal laser scanning microscopy. Results A significant loss of talin, vinculin, β-dystroglycan, β1-D and α7-D integrins were detected in obstructed UPJ, while a significant enhancement of β1-A and α7-A integrins was shown as compared to control UPJ. Conclusions In UPJO we observed an alternatively spliced isoform of both α and β subunits. Integrins play a crucial role in cell adhesion including cell-matrix and intracellular interactions, they are involved in cell migration, survival and differentiation and transmit signals by organizing the cytoskeleton.Expression of β1-A and α7-A integrins appears to be developmentally regulated and needs to meet the requirements of the different biochemical adhesion situations during SMC development. We observed a loss of talin, a flexible cystoskeletal protein important in the assembly of cell-matrix junctions, a lacking of vinculin, protein for control of apoptosis, and of β-dystroglycan, a transmembrane glycoprotein fundamental in connecting the intracellular cytoskeleton to extracellular matrix and able also to modulate the expression of α7-B integrin. Our data suggest that primary anomaly in UPJO might be attributed to a critical alteration of SMC cytoskeleton, promoting SMC abnormal function and apoptosis.