Angiotensin II and insulin induce growth of ciliary artery smooth muscle: effects of AT1/AT2 antagonists

PURPOSE: Abnormal growth of smooth muscle cells (SMCs) in small arteries of the eye is associated with hypertension and diabetes, and the complications that they induce. Migration and proliferation of SMCs into the intima are primary mechanisms involved in neointima formation. In aortic SMCs, angiotensin II (AII)-induced proliferation is inhibited by angiotensin type 1 (AT1) receptor antagonist. However, in small artery SMCs, in particular in the circulation of the eye, the effects of AII on migration and proliferation are unknown. METHODS: The effects of AII (10(-6) to 10(-10) M) on migration and proliferation of growth-arrested SMCs of porcine ciliary arteries were studied in the presence and absence of insulin (5 x 10(-10) M) by assaying DNA synthesis (3H-thymidine incorporation), cell number, and movement of SMCs across the membrane of a modified Boyden chamber. RESULTS: In the absence of insulin, only high concentrations (10(-6) to 10(-8) M) of AII induced DNA synthesis and increased cell number (P < 0.05); however, in the presence of insulin (5 x 10(-10) M), AII induced DNA synthesis and cell number at low concentrations (10(-10) M) and in a concentration-dependent manner (P < 0.05). In contrast to proliferation, AII induced SMC migration in a concentration-dependent manner in the absence of insulin (P < 0.05). The AT1 antagonist CGP48933 (10(-8) to 10(-12) M), but not the AT2 antagonist CGP42112 (10(-8) to 10(-12) M), inhibited AII (10(-8) M)-induced proliferation and migration in a concentration-dependent manner (P < 0.05). CONCLUSIONS: Our results suggest that AII is a potent mitogen for SMCs of ophthalmic arteries, an effect that is enhanced in the presence of insulin, and that it may be an important contributor to structural vascular changes in the ophthalmic circulation in hypertension associated with non-insulin dependent diabetes. The inhibition of AII-induced growth by an AT1 antagonist suggests that these drugs may be important therapeutic tools to prevent structural vascular changes in the ophthalmic vasculature under these conditions.     

Repair of chronic tympanic membrane perforations with fibroblast growth factor

A number of angiogenic growth factors have been shown to accelerate wound healing. Previous work has demonstrated that topical application of epidermal growth factor is effective in healing chronic tympanic membrane perforations in an animal model. Theoretically, fibroblast growth factor may result in a superior healed membrane through preferential stimulation of the fibroblasts within the middle layer of the tympanic membrane. To test this hypothesis, the effects of exogenously applied fibroblast growth factor on the chronically perforated tympanic membrane were evaluated. A buffered solution of fibroblast growth factor (25 microliters of fibroblast growth factor, 0.2 mg/ml) was administered to a Gelfoam pledget placed over chronic tympanic membrane perforations in chinchillas. Control ears were treated with Gelfoam and the buffer solution only. Complete closure of the tympanic membrane perforation was observed in 81% (13 of 16) of the fibroblast growth factor-treated ears, but in only 41% (7 of 17) of the controls (p = 0.05). Heading took place gradually, requiring an average of 4 weeks for the fibroblast growth factor-treated and 6.5 weeks for the control ears that healed. The relatively high healing rate for the control group does not imply that the pretreatment perforations were not chronic, rather there appears to be some efficacy to the control protocol of repeated applications of Gelfoam and buffer. A histologic analysis of the fibroblast growth factor-healed eardrums immediately after closure demonstrated hypertrophy of the squamous and fibrous layers of the tympanic membrane. Over time, the eardrum thinned to reach proportions similar to those of the normal tympanic membrane, including the presence of a substantial middle fibrous layer. A screening ototoxicity study revealed no structural damage to the organ of Corti after growth factor treatment. To assess the potential for systemic toxicity, blood and peripheral tissues were analyzed for radioactivity at time points during a 48-hour period after application of 25 microliters of 125I-fibroblast growth factor to the perforated tympanic membrane. More than 78% of the radioactivity remained at the application site. Given the tiny original dosage, the small fraction absorbed systemically is minuscule and highly unlikely to induce adverse effects in light of published toxicity data. On the basis of these promising safety and efficacy data in the chinchilla model, clinical trials of fibroblast growth factor in repair of chronic tympanic membrane perforations in human beings are being initiated.     

The regulation of collagen in fetal skin wounds: mRNA localization and analysis

The deposition of collagen in fetal skin wounds has been shown in several animal models. The authors used a radiolabeled RNA antisense probe, complementary to the mRNA for the alpha-1 chain of human procollagen type I, to assess regulation of this collagen species in fetal and adult rabbit wounds. Dorsal skin wounds were placed on fetal and maternal animals at the beginning of the third trimester, and were harvested 3, 5, and 7 days later. In situ RNA/RNA hybridization was performed on suitable specimens, and morphometric analysis was carried out with a computerized LECO image analyzer. Fetal wounds exhibited an inflow of mesenchymal cells that produced collagen type I at levels higher than the surrounding tissue; this activity was highest on days 3 and 5 after wounding. Adult wounds had increased fibroblast presence by day 7, producing collagen type I at levels higher than those of adjacent unwounded tissue. Morphometric analysis of the signal produced by in situ hybridization and of the number of cells producing the signal in a given field showed that fetal wounds appear to produce collagen type I by an increase in the number of cells in the area of the wound--not by induction of the gene for procollagen type I. In contrast, adult wounds had both fibroblast migration and induction of procollagen type I mRNA synthesis. These findings imply multilevel regulation of collagen production in the adult and posttranslational regulation in the fetus.     

Triploidy: antenatal sonographic features with post-mortem correlation

The CD34 antigen is expressed by human hematopoietic progenitor and stem cells. These cells are capable of reconstituting marrow function after marrow-ablative chemo-radiotherapy. Several different technologies have been developed for the separation of CD34+ cells from bone marrow or peripheral blood stem cell (PBSC) components. We used an immunomagnetic separation technique to enrich CD34+ cells from PBSC components in anticipation of autologous transplantation for patients with B lymphoid malignancies. Twenty-nine patients enrolled on this study and received mobilization chemotherapy followed by G-CSF. Of these, 21 achieved a peripheral blood CD34+ cell level of at least 2.0 x 10(4)/l required by protocol for separation of the stem cell components. A median of three components per patient was collected for processing. The average CD34+ cell concentration in the components after apheresis was 1.0 +/- 1.2%. After the CD34+ cell selection, the enriched components contained 0.6 +/- 0.6% of the starting nucleated cells. The recovery of CD34+ cells, however, averaged 58.4 +/- 19.2% of the starting cell number, with a purity of 90.8 +/- 6.5%. Overall depletion of CD34- cells was 99.96 +/- 0.06%. Nineteen patients were treated with marrow-ablative conditioning regimens and received an average of 6.2 +/- 2.0 x 10(6) CD34+ cells/kg body weight. These patients recovered to an ANC >0.5 x 10(9)/l at a median of 11 days (range 8-14), and platelet transfusion independence at a median of 9 days (range 5-13). Four patients died of transplant-related complications or relapse before 100 days after transplantation. No patient required infusion of unseparated cells because of failure of sustained bone marrow function. These data demonstrate that peripheral blood-derived CD34+ cells enriched by use of an immunomagnetic separation technique are capable of rapid engraftment after autologous transplantation.     

Treatment of intrahepatic cancers with radiation doses based on a normal tissue complication probability model

PURPOSE: To attempt to safely escalate the dose of radiation for patients with intrahepatic cancer, we designed a protocol in which each patient received the maximum possible dose while being subjected to a 10% risk of radiation-induced liver disease (RILD, or radiation hepatitis) based on a normal tissue complication probability (NTCP) model. We had two hypotheses: H1; with this approach, we could safely deliver higher doses of radiation than we would have prescribed based on our previous protocol, and H2; the model would predict the observed complication probability (10%). PATIENTS AND METHODS: Patients with either primary hepatobiliary cancer or colorectal cancer metastatic to the liver and normal liver function were eligible. We used an NTCP model with parameters calculated from our previous patient data to prescribe a dose that subjected each patient to a 10% complication risk within the model. Treatment was delivered with concurrent hepatic arterial fluorodeoxyuridine (HA FUdR). Patients were evaluated for RILD 2 and 4 months after the completion of treatment. RESULTS: Twenty-one patients completed treatment and were followed up for at least 3 months. The mean dose delivered by the current protocol was 56.6 +/- 2.31 Gy (range, 40.5 to 81 Gy). This dose was significantly greater than the dose that would have been prescribed by the previous protocol (46.0 +/- 1.65 Gy; range, 33 to 66 Gy; P < .01). These data are consistent with H1. One of 21 patients developed RILD. The complication rate of 4.8% (95% confidence interval, 0% to 23.8%) did not differ significantly from the predicted 8.8% NTCP (based on dose delivered) and excluded a 25% true incidence rate (P < .05). This finding supports H2. CONCLUSION: Our results suggest that an NTCP model can be used prospectively to safely deliver far greater doses of radiation for patients with intrahepatic cancer than with previous approaches. Although the observed complication probability is within the confidence intervals of our model, it is possible that this model overestimates the risk of complication and that further dose escalation will be possible. Additional follow-up and accrual will be required to determine if these higher doses produce further improvements in response and survival.     

Directed forgetting of trauma cues in adult survivors of childhood sexual abuse with and without posttraumatic stress disorder

The authors used a directed-forgetting task to investigate whether psychiatrically impaired adult survivors of childhood sexual abuse exhibit an avoidant encoding style and impaired memory for trauma cues. The authors tested women with abuse histories, either with or without posttraumatic stress disorder (PTSD), and women with neither abuse histories nor PTSD. The women saw intermixed trauma words (e.g., molested), positive words (e.g., confident), and categorized neutral words (e.g., mailbox) on a computer screen and were instructed either to remember or to forget each word. Relative to the other groups, the PTSD group did not exhibit recall deficits for trauma-related to-be-remembered words, nor did they recall fewer trauma-related to-be-forgotten words than other words. Instead, they exhibited recall deficits for positive and neutral words they were supposed to remember. These data are inconsistent with the hypothesis that impaired survivors exhibit avoidant encoding and impaired memory for traumatic information.