Veronica Triaca

We have examined the mechanisms of cell death induced by cisplatin in primary cultures of mouse proximal tubular cells. High concentrations of cisplatin (800 microM) led to necrotic cell death over a few hours. Much lower concentrations of cisplatin (8 microM) led to apoptosis, which caused loss of the cell monolayer over several days. Necrosis was characterized by a cytosolic swelling and early loss of plasma membrane integrity. In contrast, early features of cells undergoing apoptosis included cell shrinkage and loss of attachment to the monolayers. Nuclear chromatin became condensed and fragmented in apoptosing cells. These features were absent in necrotic cells. DNA electrophoresis of cells exposed to 800 microM cisplatin yielded a "smear" pattern, due to random DNA degradation. In contrast, the DNA of apoptosing cells demonstrated a "ladder" pattern resulting from internucleosomal DNA cleavage. Antioxidants delayed cisplatin-induced apoptosis but not necrosis. Thus the mechanism of cell death induced by cisplatin is concentration dependent. Reactive oxygen species play a role in mediating apoptosis but not necrosis induced by cisplatin.

We have examined the role of reactive oxygen species (ROS) in apoptosis induced by growth factor deprivation in primary cultures of mouse proximal tubular (MPT) cells. When confluent monolayers of MPT cells are deprived of all growth factors, the cells die by apoptosis over a 10- and 14-day period. Both epidermal growth factor (EGF) and high-dose insulin directly inhibit apoptosis of MPT cells deprived of growth factors. Growth factor deprivation results in an increase in the cellular levels of superoxide anion while apoptosis of MPT cells induced by growth factor withdrawal is inhibited by a number of antioxidants and scavengers of ROS. Growth factor deprivation also results in activation of caspase activity, which is inhibited by EGF and high-dose insulin as well as by the ROS scavengers and antioxidants that inhibit apoptosis. The cell-permeant caspase inhibitor, z-Val-Ala-Asp-CH2F (zVAD-fmk), prevents the increase in caspase activity and markedly inhibits apoptosis induced by growth factor deprivation. However, zVAD-fmk had no effect on the increased levels of superoxide associated with growth factor deprivation. Thus we provide novel evidence that ROS play an important role in mediating apoptosis associated with growth factor deprivation. ROS appear to act upstream of caspases in the apoptotic pathway. We hypothesize that oxidant stress, induced by growth factor withdrawal, represents a signaling mechanism for the default pathway of apoptosis.

Lysophosphatidic acid (LPA) is the smallest and structurally simplest of all glycerophospholipids. LPA is a normal constituent of serum and binds with high affinity to albumin while retaining its biological activity. The effects of LPA are pleiotropic and range from mitogenesis to stress fiber formation. In this report, we demonstrate two novel functions for LPA. LPA acts as a survival factor to inhibit apoptosis of primary cultures of mouse renal proximal tubular (MPT) cells. LPA also acts as a potent mitogen for MPT cells. The ability of LPA to act as both a survival factor and a mitogen is mediated by the lipid kinase phosphatidylinositol 3-kinase (PI3K), since these activities were completely blocked by wortmannin or LY-294002, two structurally dissimilar inhibitors of PI3K. The identification of LPA as a proliferative and anti-apoptotic factor suggests a potential role for this lipid mediator during the injury and/or recovery phases following tubular damage.

PURPOSE: We evaluated the efficacy of a combined chemoradiation therapy protocol for the primary treatment of primary invasive carcinoma of the male urethra. MATERIALS AND METHODS: From January 1991 to December 2006, 18 patients with invasive carcinoma of the male urethra referred to our institution were treated with a chemoradiation therapy protocol, consisting of 2 cycles of 5-fluorouracil (1,000 mg/m(2)) on days 1 to 4 and days 29 to 32, and mitomycin-C (10 mg/m(2)) on days 1 and 29 with concurrent external beam radiation therapy (45 to 55 Gy in 25 fractions during 5 weeks) to the genitalia, perineum, and inguinal and external iliac lymph nodes. Kaplan-Meier curves were constructed to assess overall, disease specific and disease-free survival. RESULTS: The stage and node distribution was T2N0 in 2 patients (11%), T3N0 in 8 (44%), T4N0 in 2 (11%), TXN1 in 1(6%) and TXN2 in 5 (28%). The most prevalent histology was moderately (7 of 18 patients or 39%) or poorly (10 of 18 or 56%) differentiated squamous cell carcinoma (17 of 18 or 95%). Overall 83% (15 of 18) of the patients had a complete response to the primary chemoradiation therapy protocol, and the 5-year overall and disease specific survival rates were 60% and 83%, respectively. Five-year disease-free survival rates after chemoradiation therapy and after chemoradiation therapy with salvage surgery were 54% and 72%, respectively. The 3 nonresponders died of disease after undergoing salvage surgery and 5 of the 15 complete responders (30%) had recurrence. Complex urethral reconstruction was required in 3 of 10 patients (30%) who had prolonged disease-free survival. CONCLUSIONS: The chemoradiation therapy protocol is an alternative primary treatment modality for invasive urethral carcinoma. It enables an unprecedented potential for organ preservation.

PURPOSE OF REVIEW: To summarize recent evidence suggesting a genetic basis for the development of urogenital prolapse and stress urinary incontinence. RECENT FINDINGS: Epidemiological evidence suggests that some women have a genetic predisposition to the development of urogenital prolapse and stress incontinence. Abnormal expression of various structural proteins is thought to be the molecular genetic mechanism for the development of these conditions. A group of families with an autosomal dominant pattern of transmission of urogenital prolapse with high penetrance has been identified. No similar cohort of families with familial stress incontinence currently exists, although candidate genes have been identified that appear to predispose women to urogenital prolapse and stress incontinence. Additionally, animal models of urogenital prolapse have been developed that closely parallel the development of prolapse in humans. SUMMARY: A growing body of evidence suggests a genetic basis for the development of urogenital prolapse and stress incontinence. Candidate genes have been identified that may result in alteration of the normal metabolism of various structural proteins which may ultimately predispose some women to both urogenital prolapse and stress incontinence. Further research into the genetic basis of these conditions may provide a comprehensive understanding of the biological basis of these disorders.