Complications of single-port robot-assisted radical prostatectomy: multi-institutional analysis from the Single-Port Advanced Research Consortium (SPARC).

OBJECTIVE: To evaluate the perioperative complications of single-port robot-assisted radical prostatectomy (SP-RARP). PATIENTS AND METHODS: A retrospective review was performed on the prospectively maintained, Institutional Review Board-approved, multi-institutional Single-Port Advanced Research Consortium (SPARC) database. A total of 1103 patients were identified who underwent three different approaches of SP-RARP between 2019 and 2022 using the purpose-built SP robotic platform. In addition to baseline clinical, perioperative outcomes, this study comprehensively analysed for any evidence of intraoperative complication, as well as postoperative complication and readmission within 90 days of the respective surgery. RESULTS: Of the 244, 712, and 147 patients who underwent transperitoneal, extraperitoneal, and transvesical SP-RARP, respectively, intraoperative complications were noted in five patients (0.4%), all of which occurred during the transperitoneal approach. Two patients had bowel serosal tears, two had posterior button-holing of the bladder necessitating repair, and one patient had an obturator nerve injury. Postoperative complications were noted in 143 patients (13%) with major complications (Clavien-Dindo Grade ≥III) only identified in 3.7% of the total cohort. The most common complications were lymphocele (3.9%), acute urinary retention (2%), and urinary tract infection (1.9%). The 90-day re-admission rate was 3.9%. CONCLUSION: The SP-RARP is a safe and effective procedure with low complication and readmission rates regardless of the approach. These results are comparable to current multi-port RARP literature.

Low Risk of Postoperative Hernia Following Single-port Robot-assisted Radical Prostatectomy: A Report From the Single-port Advanced Research Consortium (SPARC).

OBJECTIVE: To evaluate the risk of postoperative hernia following different approaches of single-port robot-assisted radical prostatectomy (SP-RARP). METHODS: A retrospective review was performed on patients who underwent SP-RARP between February 2019 and December 2022. Demographic and clinical information was collected from the multi-institutional, prospectively-maintained Single-Port Advanced Research Consortium (SPARC) database. Data were analyzed using IBM Statistical Packaging for Social Sciences (SPSS) version 29.0 with descriptive statistics as presented. RESULTS: A total of 1103 patients were identified, consisting of 244 (22.1%), 712 (64.6%), and 147 (13.3%) cases performed via transperitoneal, extraperitoneal (EP), and transvesical (TV) approaches, respectively. During a median follow-up time of 11 months (interquartile range 5.7-17.1 months), only two cases of incisional hernia were reported. Both cases occurred following transperitoneal SP-RARP with one patient requiring surgical repair. There remains no evidence of postoperative hernia following EP and TV SP-RARP at the completion of our review. CONCLUSION: SP-RARP was associated with low risk for postoperative hernia. The risk was lower following TV and EP SP-RARP where the peritoneum is preserved.

Single-port Extraperitoneal and Transperitoneal Radical Prostatectomy: A Multi-Institutional Propensity-Score Matched Study.

OBJECTIVE: To compare the perioperative and early postoperative outcomes between single-port (SP) extraperitoneal radical prostatectomy (EPRP) and SP transperitoneal radical prostatectomy (TPRP), in a multi-institutional setting. METHODS: We identified all patients who underwent SP robot-assisted radical prostatectomy at 6 different institutes. Data of 650 patients were collected and divided into 2 groups based on the surgical approach: SP EPRP or SP TPRP. A Propensity-score matched-pair analysis for body mass index (BMI), prostate size, and National Comprehensive Cancer Network risk was performed with a 1:1 ratio. Analysis of perioperative and postoperative outcomes was performed using Wilcoxon signed-rank test and chi-square and Fisher's exact tests. RESULTS: After matching, 238 patients were included in each arm. The median follow-up period was 7 and 6 months for EPRP and TPRP groups, respectively. The total operative time was longer in the EPRP group (206 vs 155 minutes, P < .001). The EPRP group had a shorter length of hospitalization and same-day discharge rate compared to the TPRP approach (P < .001). There was no difference in the overall intraoperative or postoperative complications rate between the 2 groups, nor positive surgical margin rates. CONCLUSION: The SP extraperitoneal approach is associated with a shorter hospital stay and higher rate of same-day discharge, with no difference in the surgical margin, or complication rates.

Single-Port Robotic Radical Prostatectomy: Short-Term Outcomes and Learning Curve.

Introduction and Objective: In 2018, the U.S. Food and Drug Administration approved the da Vinci single-port (SP) system, in which four instruments are still utilized, but enter through a single-site access trocar. Herein, we report the largest case series for SP robot-assisted radical prostatectomy (RARP) to date. Our primary aim is to analyze the perioperative and short-term outcomes of this procedure. Our secondary aim is an assessment of the learning curve with this new platform. Methods: A total of 157 patients underwent SP RARP by two surgeons who have completed >3000 multiport robotic surgeries collectively. Institutional Review Board-approved prospectively collected data were used. Basic demographic preoperative variables and perioperative outcomes were analyzed. Results: Median patient age and prostate-specific antigen was 63 years and 6.3 ng/mL before treatment (interquartile range [IQR] 4.7-8.2 ng/mL). Average prostate weight was 47 g. The median operating time was 195 minutes (IQR 165-221.25 minutes) with a median estimated blood loss of 100 mL (IQR 100-200 mL). Surgeon 1's operating time stabilized around case #56, and Surgeon 2 around case #26. Surgeon 2 used the transperitoneal approach for the first 7 cases. There were no intraoperative complications. There were six total postoperative complications (3.8%) and four (2.5%) were Clavien-Dindo scale ≥IIIa. One hundred ten patients went home same day, 45 stayed 1 night at the hospital, with only 2 patients requiring stay in the hospital for more than 1 night (70%, 29%, and 1% respectively). With the median follow-up period of 9 months, rates of biochemical recurrence, pad-free, and potency preservation were 8.3%, 82.5%, and 64.4%, respectively. Conclusions: This case series confirms the safety and efficacy of SP RARP with acceptable short-term outcomes. There is a significant learning curve for this new modality. Shorter hospital stay appears to be an early benefit of the SP platform.

In vivo fluorescence imaging of conjunctival goblet cells.

Conjunctival goblet cells (GCs) are specialized epithelial cells that secrete mucins onto the ocular surface to maintain the wet environment. Assessment of GCs is important because various ocular surface diseases are associated with their loss. Although there are GC assessment methods available, the current methods are either invasive or difficult to use. In this report, we developed a simple and non-invasive GC assessment method based on fluorescence imaging. Moxifloxacin ophthalmic solution was used to label GCs via topical administration, and then various fluorescence microscopies could image GCs in high contrasts. Fluorescence imaging of GCs in the mouse conjunctiva was confirmed by both confocal reflection microscopy and histology with Periodic acid-Schiff (PAS) labeling. Real-time in-vivo conjunctival GC imaging was demonstrated in a rat model by using both confocal fluorescence microscopy and simple wide-field fluorescence microscopy. Different GC densities were observed in the forniceal and bulbar conjunctivas of the rat eye. Moxifloxacin based fluorescence imaging provides high-contrast images of conjunctival GCs non-invasively and could be useful for the study or diagnosis of GC related ocular surface diseases.

Comparison of Reading Speed after Bilateral Bifocal and Trifocal Intraocular Lens Implantation.

PURPOSE: To evaluate and compare visual acuity and reading speed for Korean language between a diffractive bifocal and trifocal intraocular lens (IOL) of the same material and haptic design. METHODS: We reviewed the medical records of the patients who had undergone bilateral cataract surgery with bifocal IOLs (AT LISA 801) on the both eyes (bifocal group) and trifocal IOLs (AT LISA tri 839 MP, trifocal group). The main outcome measures were the uncorrected distance, intermediate, and near visual acuity (uncorrected distance visual acuity [UCDVA], uncorrected intermediate visual acuity [UCIVA], and uncorrected near visual acuity [UCNVA]) and corrected distance, near, and distance-corrected intermediate visual acuity (corrected distance visual acuity [CDVA], corrected near visual acuity [CNVA], and distance-corrected intermediate visual acuity [DCIVA]) at last postoperative follow-up month. Reading speeds for Korean language were measured to check near visual function. RESULTS: Fourteen eyes (7 patients) were included in the bifocal group and 32 eyes of 16 patients in the trifocal group. There were no statistical differences between the two groups with respect to UCDVA, UCNVA, CDVA, and CNVA. However, UCIVA (0.35 vs. 0.22 logarithm of the minimum angle of resolution [logMAR], p < 0.01) and DCIVA (0.34 vs. 0.20 logMAR, p < 0.01) were significantly better in the trifocal group than in the bifocal group. The mean reading speed for logMAR 0.5 optotype (point 10) was 86.50 words per minute (wpm) in the bifocal group and 81.48 wpm in the trifocal group without a significant difference (p = 0.70). CONCLUSIONS: Trifocal IOLs provided the same level of distance and near visual acuity and reading speed as that of bifocal IOLs with better intermediate visual acuity.

Implementation of a bundled protocol significantly reduces risk of spinal cord ischemia after branched or fenestrated endovascular aortic repair.

OBJECTIVE: Spinal cord ischemia (SCI) is a devastating complication after branched or fenestrated endovascular aortic repair (B/FEVAR) for thoracoabdominal aortic disease. The purpose of this analysis was to describe the impact of a bundled clinical care protocol designed to reduce the risk of SCI in this population of patients. METHODS: A bundled SCI prevention protocol including cerebrospinal fluid drainage, blood pressure parameters, transfusion goals, and pharmacologic adjuncts (steroids, naloxone) was initiated in May 2015. Before that date, portions of the protocol (cerebrospinal fluid drainage in particular) were used in an informal fashion in patients perceived to be at high risk. B/FEVAR cases completed from January 2012 to May 2016 were reviewed, and outcomes before (n = 223) and after (n = 70) SCI bundle application were compared. The primary end point was the incidence of SCI events. Secondary end points included length of stay, complications, and survival. High-risk patients for SCI were defined as those undergoing B/FEVAR resulting in aortic coverage equivalent to open Crawford extent I to III thoracoabdominal aortic aneurysm (TAAA) repair. Survival was estimated using Kaplan-Meier life-table analysis. RESULTS: Postprotocol patients were more likely to be older (75 ± 7 vs 72 ± 8 years; P = .03), to have an American Society of Anesthesiologists class 4 designation (94% vs 81%; P = .04), and to be treated for TAAA (67% vs 56%; P = .004). Postprotocol pre-emptive spinal drain use was greater in high-risk patients (100% vs 87%; P = .04) but significantly decreased in lower risk patients (suprarenal aneurysm or extent IV TAAA: 5% after protocol implementation vs 21% before protocol implementation; P = .04). Rates of any SCI before and after implementation of the bundled protocol were 13% (n = 29 of 223) and 3% (n = 2 of 70; P = .007), respectively. In comparing high-risk patients, protocol use resulted in an even more significant reduction in SCI rate (19% [28 of 144] vs 4% [2 of 50]; P = .004). Postoperative morbidity (41% vs 33%; P = .2) and 30-day mortality (5% vs 1%; P = .3) were not different between groups. However, patients treated on protocol had significantly improved 1-year survival (99% ± 1% after protocol implementation vs 90% ± 2% before protocol implementation; log-rank, P = .05). CONCLUSIONS: Implementation of a bundled multimodal protocol may significantly reduce risk of SCI after B/FEVAR, with the greatest risk reduction occurring in the most vulnerable patients. Interestingly, reduction in SCI risk was associated with improvement in 1-year survival.

Korean Version Self-testing Application for Reading Speed.

PURPOSE: This study introduces a reading chart application for the iPad tablet in the Korean language and investigates the reading speed in a normal-sighted population according to age group. METHODS: Sixty-three Korean sentences were selected from textbooks for second grade elementary school students. A commonly used typeface in everyday printed material, "BatangChe," was used. Letter size was presented in logarithm of the minimum angle of resolution (logMAR) 0.0 to 1.0 at 0.1 logMAR steps at a reading distance of 40 cm. A third generation retina display iPad was used to present the chart, and the sentences were presented randomly for each size of letter. The subjects repeated the test silently (reading only) and out loud (reading and speaking) to prevent them from skipping reading words. Pilot testing followed in 65 normal vision adults under 60 years of age. RESULTS: The mean reading only speed for logMAR 0.5 optotype (point 10) was 121.1 ± 47.2 words per minute (wpm) for people in their 20s (n=21), 116.5 ± 38.3 in their 30s (n=27), 93.8 ± 12.6 in their 40s (n=9), and 56.5 ± 42.7 (n=8) in their 50s. There was a significant correlation between age and reading and speaking speed (r=-0.48, p<0.001). The mean reading only speed for logMAR 0.5 optotype (point 10) was 202.3 ± 88.4 wpm and the mean reading and speaking speed was 129.7 ± 25.9 wpm, with significantly different (p<0.001). CONCLUSIONS: This Korean reading chart application could present a new standard when checking reading speed according to age groups.

Pseudomonas aeruginosa wound infection involves activation of its iron acquisition system in response to fascial contact.

BACKGROUND: Wound infections are traditionally thought to occur when microbial burden exceeds the innate clearance capacity of host immune system. Here, we introduce the idea that the wound environment itself plays a significant contributory role to wound infection. METHODS: We developed a clinically relevant murine model of soft tissue infection to explore the role of activation of microbial virulence in response to tissue factors as a mechanism by which pathogenic bacteria cause wound infections. Mice underwent abdominal skin incision and light muscle injury with a crushing forceps versus skin incision alone followed by topical inoculation of Pseudomonas aeruginosa. Mice were sacrificed on postoperative Day 6, and abdominal tissues were analyzed for clinical signs of wound infection. To determine if specific wound tissue components induce bacterial virulence, P. aeruginosa was exposed to the skin, fascia, and muscle. RESULTS: Gross wound infection caused by P. aeruginosa was observed to be significantly increased in injured tissues versus noninjured (80% vs.10%) tissues (n = 20 per group, p < 0.0001). Exposure of P. aeruginosa to individual tissue components demonstrated that fascia significantly induced bacterial virulence as judged by the production of pyocyanin, a redox-active phenazine compound known to kill immune cells. Whole-genome transcriptional profiling of P. aeruginosa exposed to the fascia demonstrated activation of multiple genes responsible for the synthesis of the iron scavenging molecule pyochelin. CONCLUSION: We conclude that wound elements, in particular fascia, may play a significant role in enhancing the virulence of P. aeruginosa and may contribute to the pathogenesis of clinical wound infection.

The intestinal environment of surgical injury transforms Pseudomonas aeruginosa into a discrete hypervirulent morphotype capable of causing lethal peritonitis.

BACKGROUND: Secondary peritonitis continues to carry a high mortality rate despite the aggressive use of imaging, drainage, and antibiotics. Although host factors and microbial burden contribute to the outcome of peritonitis, we propose a role for bacterial virulence as a determinant of outcome from peritonitis. Bacterial virulence is an inducible trait that is activated in response to specific local "cues" that we have previously shown to be present in the mouse gut exposed to surgical stress and injury. METHODS: Pseudomonas aeruginosa was harvested after its intestinal inoculation into the cecum of mice subjected to surgical injury (30% hepatectomy) or sham surgery (controls). Harvested strains were then injected into the peritoneum of noninjured (naïve) mice and mortality determined. RESULTS: P. aeruginosa harvested from the intestines of surgically injured mice caused 100% mortality, whereas strains harvested from control mice caused no mortality. Among recovered strains, a distinct P. aeruginosa morphotype (wrinkled shape) was shown to cause lethal peritonitis compared to smooth-shaped strains, which were nonlethal. Wrinkled strains were associated with a tendency to elicit a more proinflammatory response in mice compared to smooth-shaped strains. CONCLUSION: Surgical injury transforms the morphotype of intestinal P. aeruginosa to express a hypervirulent response in the peritoneum of mice. Enhanced virulence of intestinal pathogens in response to surgical injury may play an important role in predicting the outcome of peritonitis.

Using an Agent-Based Model to Examine the Role of Dynamic Bacterial Virulence Potential in the Pathogenesis of Surgical Site Infection.

OBJECTIVE: Despite clinical advances, surgical site infections (SSIs) remain a problem. The development of SSIs involves a complex interplay between the cellular and molecular mechanisms of wound healing and contaminating bacteria, and here, we utilize an agent-based model (ABM) to investigate the role of bacterial virulence potential in the pathogenesis of SSI. APPROACH: The Muscle Wound ABM (MWABM) incorporates muscle cells, neutrophils, macrophages, myoblasts, abstracted blood vessels, and avirulent/virulent bacteria to simulate the pathogenesis of SSIs. Simulated bacteria with virulence potential can mutate to possess resistance to reactive oxygen species and increased invasiveness. Simulated experiments (t=7 days) involved parameter sweeps of initial wound size to identify transition zones between healed and nonhealed wounds/SSIs, and to evaluate the effect of avirulent/virulent bacteria. RESULTS: The MWABM reproduced the dynamics of normal successful healing, including a transition zone in initial wound size beyond which healing was significantly impaired. Parameter sweeps with avirulent bacteria demonstrated that smaller wound sizes were associated with healing failure. This effect was even more pronounced with the addition of virulence potential to the contaminating bacteria. INNOVATION: The MWABM integrates the myriad factors involved in the healing of a normal wound and the pathogenesis of SSIs. This type of model can serve as a useful framework into which more detailed mechanistic knowledge can be embedded. CONCLUSION: Future work will involve more comprehensive representation of host factors, and especially the ability of those host factors to activate virulence potential in the microbes involved.

Immature oxidative stress management as a unifying principle in the pathogenesis of necrotizing enterocolitis: insights from an agent-based model.

BACKGROUND: Necrotizing enterocolitis (NEC) is a complex disease involving prematurity, enteral feeding, and bacterial effects. We propose that the underlying initial condition in its pathogenesis is reduced ability of the neonatal gut epithelial cells (NGECs) to clear oxidative stress (OS), and that when such a NGEC population is exposed to enteral feeding, the increased metabolic OS tips the population toward apoptosis, inflammation, bacterial activation, and eventual necrosis. The multi-factorial complexity of NEC requires characterization with computational modeling, and herein, we used an agent-based model (ABM) to instantiate and examine our unifying hypothesis of the pathogenesis of NEC. METHODS: An ABM of the neonatal gut was created with NGEC computational agents incorporating rules for pathways for OS, p53, tight junctions, Toll-like receptor (TLR)-4, nitric oxide, and nuclear factor-kappa beta (NF-κB). The modeled bacteria activated TLR-4 on contact with NGECs. Simulations included parameter sweeps of OS response, response to feeding, addition of bacteria, and alterations in gut mucus production. RESULTS: The ABM reproduced baseline cellular respiration and clearance of OS. Reduction in OS clearance consistent with clinical NEC led to senescence, apoptosis, or inflammation, with disruption of tight junctions, but rarely to NGEC necrosis. An additional "hit" of bacteria activating TLR-4 potentiated a shift to NGEC necrosis across the entire population. The mucus layer was modeled to limit bacterial-NGEC interactions and reduce this effect, but concomitant apoptosis in the goblet cell population reduced the efficacy of the mucus layer and limited its protective effect in simulated experiments. This finding suggests a means by which increased apoptosis at the cellular population level can lead to a transition to the necrosis outcome. CONCLUSIONS: Our ABM incorporates known components of NEC and demonstrates that impaired OS management can lead to apoptosis and inflammation of NGECs, rendering the system susceptible to an additional insult involving regionalized mucus barrier failure and TLR-4 activation, which potentiates the necrosis outcome. This type of integrative dynamic knowledge representation can be a useful adjunct to help guide and contextualize research.

Pseudomonas aeruginosa potentiates the lethal effect of intestinal ischemia-reperfusion injury: the role of in vivo virulence activation.

BACKGROUND: Experimental models of intestinal ischemia-reperfusion (IIR) injury are invariably performed in mice harboring their normal commensal flora, even though multiple IIR events occur in humans during prolonged intensive care confinement when they are colonized by a highly pathogenic hospital flora. The aims of this study were to determine whether the presence of the human pathogen Pseudomonas aeruginosa in the distal intestine potentiates the lethality of mice exposed to IIR and to determine what role any in vivo virulence activation plays in the observed mortality. METHODS: Seven- to 9-week-old C57/BL6 mice were exposed to 15 minutes of superior mesenteric artery occlusion (SMAO) followed by direct intestinal inoculation of 1.0 × 10(6) colony-forming unit of P. aeruginosa PAO1 into the ileum and observed for mortality. Reiterative studies were performed in separate groups of mice to evaluate both the migration/dissemination pattern and in vivo virulence activation of intestinally inoculated strains using live photon camera imaging of both a constitutive bioluminescent P. aeruginosa PAO1 derivative XEN41 and an inducible reporter derivative of PAO1, the PAO1/lecA:luxCDABE that conditionally expresses the quorum sensing-dependent epithelial disrupting virulence protein PA 1 Lectin (PA-IL). RESULTS: Mice exposed to 15 minutes of SMAO and reperfusion with intestinal inoculation of P. aeruginosa had a significantly increased mortality rate (p < 0.001) of 100% compared with <10% for sham-operated mice intestinally inoculated with P. aeruginosa without SMAO and IIR alone (<50%). Migration/dissemination patterns of P. aeruginosa in mice subjected to IIR demonstrated proximal migration of distally injected strains and translocation to mesenteric lymph nodes, liver, spleen, lung, and kidney. A key role for in vivo virulence expression of the barrier disrupting adhesin PA-IL during IIR was established since its expression was enhanced during IR and mutant strains lacking PA-IL displayed attenuated mortality. CONCLUSIONS: The presence of intestinal P. aeruginosa potentiates the lethal effect of IIR in mice in part due to in vivo virulence activation of its epithelial barrier disrupting protein PA-IL.

Robot-assisted laparoscopic excision of a pelvic extragastrointestinal stromal tumor: a case report and literature review.

A 61-year-old male presented with long standing urinary frequency and the sensation of incomplete emptying. Computed tomography (CT) revealed a 9.5 cm x 7.9 cm x 6.9 cm pelvic mass behind the bladder and abutting the rectum. The mass was excised using a robotic-assisted laparoscopic approach. Pathologic examination of the mass demonstrated an extragastrointestinal stromal tumor (EGIST), an extremely rare entity. To the best of our knowledge, this is the first EGIST to be found in the rectovesicular pouch of a male and the first to be resected robotically. Our case adds to the understanding of EGISTs and their possible origin and demonstrates that robotic-assisted resection of large pelvic masses can be safe and potentially curative.

Transforming growth factor-beta : biology and clinical relevance.

Transforming growth factor-beta is a pleiotropic growth factor that has enthralled many investigators for approximately two decades. In addition to many reports that have clarified the basic mechanism of transforming growth factor-beta signal transduction, numerous laboratories have published on the clinical implication/application of transforming growth factor-beta . To name a few, dysregulation of transforming growth factor-beta signaling plays a role in carcinogenesis, autoimmunity, angiogenesis, and wound healing. In this report, we will review these clinical implications of transforming growth factor-beta .

Restoration of bone morphogenetic protein receptor type II expression leads to a decreased rate of tumor growth in bladder transitional cell carcinoma cell line TSU-Pr1.

Bone morphogenetic proteins (BMPs), potential regulators of cellular growth and metastasis that signal through an interaction with plasma membrane receptors, have been suggested to be important regulators of malignant cells. The present study was carried out to evaluate the potential role of BMP receptor (BMP-R) types IA, IB, and II in bladder transitional cell carcinoma (TCC) cells. Initially, we investigated the expression of these BMP-Rs in 30 archival tissues of human bladder TCC using immunohistochemistry; 10 benign bladder specimens were used for comparison. The results demonstrated that the expression of BMP-Rs is localized preferentially to the transitional epithelium and that there was a significant association between loss of BMP-RII expression and tumor grade. To find a cell line that can serve as a model system for clinical observation, we subsequently examined sensitivity to BMP-4 and expression of BMP-RII, BMP-RIA, and BMP-RIB in three human bladder cancer cell lines, TCC-Sup, RT4, and TSU-Pr1. Of the three cell lines, TSU-Pr1 exhibited a decreased level of BMP-RII expression and was resistant to the growth-inhibitory effect of BMP-4. Overexpression of BMP-RII in TSU-Pr1 cells not only restored BMP-4 responsiveness but also significantly decreased tumorigenicity in vivo. Taken together, these results demonstrate that human bladder TCC tissues have a frequent loss of BMP-RII expression and that overexpression of BMP-RII leads to restoration of BMP signaling and decreased tumor growth in the human bladder TCC cell line TSU-Pr1.

Loss of expression of bone morphogenetic protein receptor type II in human prostate cancer cells.

Bone morphogenetic proteins (BMPs) are members of the transforming growth factor-beta superfamily and signal through a number of membrane receptors. We have previously demonstrated that the loss of expression of BMP receptors (BMPRs) type IA, -IB, and -II (BMP-RIA, -RIB, and -RII) correlates with Gleason score in prostate cancer patients. To evaluate the prognostic value of this observation, we used immunohistochemistry to investigate the expression of BMPRs in association with disease progression in 60 patients. The results demonstrated a significant association between the loss of expression of the three BMPRs and Gleason score and clinical stage. However, only the loss of expression of BMP-RII showed a statistically significant association with 5-year survival rate (P<0.05) and biochemical recurrence-free rate following radical prostatectomy (P<0.005). To elucidate the effect of an abnormal BMP signaling in prostate cancer cells, we transfected dominant-negative BMP-RII (BMP-RIIDN) into the human prostate cancer cell line, PC3M. When a stable clone overexpressing BMP-RIIDN was inoculated subcutaneously into nude mice, the tumor growth rate was approximately 10 times that of control and parental cell line. These observations, taken together, indicate that the loss of BMP-RII expression as measured by immunohistochemistry may be a prognostic marker in prostate cancer patients, and that the loss of BMP-RII function may result in increased tumorigenicity in human prostate cancer cells.

Catalytically active human telomerase mutants with allele-specific biological properties.

Expression of the catalytic subunit of human telomerase, hTERT, extends human primary fibroblast life span. Such life span extension has generally been reported to be accompanied by net telomere lengthening, which led to the hypothesis that it is the telomere lengthening that causes the life span extension. Here we show that hTERT+C and hTERT-FlagC, mutant telomerase proteins with either 10 additional residues or a FLAG epitope added to the hTERT C-terminus, confer significant but limited life span extension to IMR90 human primary lung fibroblasts. However, as the cells continue to grow for >100 population doublings past their normal senescence point, bulk telomere length continues to erode to lengths much shorter than those seen at the senescence of control telomerase-negative cells. Expression of hTERT+C immortalized IMR90 cells transformed by three different oncogenes. Again, bulk telomeres became much shorter than those of the control cells at crisis. Additional hTERT mutants were constructed and analyzed similarly. Enzymatically active hTERT-N125A+T126A, like other previously reported conserved GQ domain mutants and C-terminally HA-tagged hTERT, failed to extend life span. Another GQ domain mutant, hTERT-E79A, was indistinguishable from wild-type hTERT in its cell growth effects, but there was no net telomere lengthening. These results uncover further hTERT allele-specific phenotypes that uncouple telomerase activity, net telomere lengthening and life span extension.