Double-Sided Sapphire Optrodes with Conductive Shielding Layers to Reduce Optogenetic Stimulation Artifacts.

Optrodes, which are single shaft neural probes integrated with microelectrodes and optical light sources, offer a remarkable opportunity to simultaneously record and modulate neural activities using light within an animal's brain; however, a common problem with optrodes is that stimulation artifacts can be observed in the neural recordings of microelectrodes when the light source on the optrode is activated. These stimulation artifacts are undesirable contaminants, and they cause interpretation complexity when analyzing the recorded neural activities. In this paper, we tried to mitigate the effects of the stimulation artifacts by developing a low-noise, double-sided optrode integrated with multiple Electromagnetic Shielding (EMS) layers. The LED and microelectrodes were constructed separately on the top epitaxial and bottom substrate layers, and EMS layers were used to separate the microelectrodes and LED to reduce signal cross-talks. Compared with conventional single-sided designs, in which the LED and microelectrodes are constructed on the same side, our results indicate that double-sided optrodes can significantly reduce the presence of stimulation artifacts. In addition, the presence of stimulation artifacts can further be reduced by decreasing the voltage difference and increasing the rise/fall time of the driving LED pulsed voltage. With all these strategies, the presence of stimulation artifacts was significantly reduced by ~76%. As well as stimulation suppression, the sapphire substrate also provided strong mechanical stiffness and support to the optrodes, as well as improved electronic stability, thus making the double-sided sapphire optrodes highly suitable for optogenetic neuroscience research on animal models.

Influence of the Surface Material and Illumination upon the Performance of a Microelectrode/Electrolyte Interface in Optogenetics.

Integrated optrodes for optogenetics have been becoming a significant tool in neuroscience through the combination of offering accurate stimulation to target cells and recording biological signals simultaneously. This makes it not just be widely used in neuroscience researches, but also have a great potential to be employed in future treatments in clinical neurological diseases. To optimize the integrated optrodes, this paper aimed to investigate the influence of surface material and illumination upon the performance of the microelectrode/electrolyte interface and build a corresponding evaluation system. In this work, an integrated planar optrode with a blue LED and microelectrodes was designed and fabricated. The charge transfer mechanism on the interface was theoretically modeled and experimentally verified. An evaluation system for assessing microelectrodes was also built up. Using this system, the proposed model of various biocompatible surface materials on microelectrodes was further investigated under different illumination conditions. The influence of illumination on the microelectrode/electrolyte interface was the cause of optical artifacts, which interfere the biological signal recording. It was found that surface materials had a great effect on the charge transfer capacity, electrical stability and recoverability, photostability, and especially optical artifacts. The metal with better charge transfer capacity and electrical stability is highly possible to have a better performance on the optical artifacts, regardless of its electrical recoverability and photostability under the illumination conditions of optogenetics. Among the five metals used in our investigation, iridium served as the best surface material for the proposed integrated optrodes. Thus, optimizing the surface material for optrodes could reduce optical interference, enhance the quality of the neural signal recording for optogenetics, and thus help to advance the research in neuroscience.

OPTICC: A multicentre trial of Occult Pneumothoraces subjected to mechanical ventilation: The final report.

INTRODUCTION: Patients with occult pneumothorax (OPTX) requiring positive-pressure ventilation (PPV) face uncertain risks of tension pneumothorax or chest drainage complications. METHODS: Adults with traumatic OPTXs requiring PPV were randomized to drainage/observation, with the primary outcome of composite "respiratory distress" (RD)). RESULTS: Seventy-five (75) patients were randomized to observation, 67 to drainage. RD occurred in 38% observed and 25% drained (p = 0.14; Power = 0.38), with no mortality differences. One-quarter of observed patients failed, reaching 40% when ventilated >5 days. Twenty-three percent randomized to drainage had complications or ineffectual drains. CONCLUSION: RD was not significantly different with observation. Thus, OPTXs may be cautiously observed in stable patients undergoing short-term PPV when prompt "rescue drainage" is immediately available. As 40% of patients undergoing prolonged (≥5 days) ventilation (PPPV) require drainage, we suggest consideration of chest drainage performed with expert guidance to reduce risk of chest tube complications. LEVEL OF EVIDENCE: Therapeutic study, level II.

The prognostic value of serum procalcitonin measurements in critically injured patients: a systematic review.

BACKGROUND: Major trauma is associated with high incidence of septic complications and multiple organ dysfunction (MOD), which markedly influence the outcome of injured patients. Early identification of patients at risk of developing posttraumatic complications is crucial to provide early treatment and improve outcomes. We sought to evaluate the prognostic value of serum procalcitonin (PCT) levels after trauma as related to severity of injury, sepsis, organ dysfunction, and mortality. METHODS: We searched PubMed, MEDLINE, EMBASE, the Cochrane Database, and references of included articles. Two investigators independently identified eligible studies and extracted data. We included original studies that assessed the prognostic value of serum PCT levels in predicting severity of injury, sepsis, organ dysfunction, and mortality among critically injured adult patients. RESULTS: Among 2015 citations, 19 studies (17 prospective; 2 retrospective) met inclusion criteria. Methodological quality of included studies was moderate. All studies showed a strong correlation between initial PCT levels and Injury Severity Score (ISS). Twelve out of 16 studies demonstrated significant elevation of initial PCT levels in patients who later developed sepsis after trauma. PCT level appeared a strong predictor of MOD in seven out of nine studies. While two studies did not show association between PCT levels and mortality, four studies demonstrated significant elevation of PCT levels in non-survivors versus survivors. One study reported that the PCT level of ≥ 5 ng/mL was associated with significantly increased mortality (OR 3.65; 95% CI 1.03-12.9; p = 0.04). CONCLUSION: PCT appears promising as a surrogate biomarker for trauma. Initial peak PCT level may be used as an early predictor of sepsis, MOD, and mortality in trauma population.

High Mobility Group Box-1 Protein and Outcomes in Critically Ill Surgical Patients Requiring Open Abdominal Management.

Background. Previous studies assessing various cytokines in the critically ill/injured have been uninformative in terms of translating to clinical care management. Animal abdominal sepsis work suggests that enhanced intraperitoneal (IP) clearance of Damage-Associated Molecular Patterns (DAMPs) improves outcome. Thus measuring the responses of DAMPs offers alternate potential insights and a representative DAMP, High Mobility Group Box-1 protein (HMGB-1), was considered. While IP biomediators are being recognized in critical illness/trauma, HMGB-1 behaviour has not been examined in open abdomen (OA) management. Methods. A modified protocol for HMGB-1 detection was used to examine plasma/IP fluid samples from 44 critically ill/injured OA patients enrolled in a randomized controlled trial comparing two negative pressure peritoneal therapies (NPPT): Active NPPT (ANPPT) and Barker's Vacuum Pack NPPT (BVP). Samples were collected and analyzed at the time of laparotomy and at 24 and 48 hours after. Results. There were no statistically significant differences in survivor versus nonsurvivor HMGB-1 plasma or IP concentrations at baseline, 24 hours, or 48 hours. However, plasma HMGB-1 levels tended to increase continuously in the BVP cohort. Conclusions. HMGB-1 appeared to behave differently between NPPT cohorts. Further studies are needed to elucidate the relationship of HMGB-1 and outcomes in septic/injured patients.

Inflammatory mediators in intra-abdominal sepsis or injury - a scoping review.

INTRODUCTION: Inflammatory and protein mediators (cytokine, chemokine, acute phase proteins) play an important, but still not completely understood, role in the morbidity and mortality of intra-abdominal sepsis/injury. We therefore systematically reviewed preclinical and clinical studies of mediators in intra-abdominal sepsis/injury in order to evaluate their ability to: (1) function as diagnostic/prognostic biomarkers; (2) serve as therapeutic targets; and (3) illuminate the pathogenesis mechanisms of sepsis or injury-related organ dysfunction. METHODS: We searched MEDLINE, PubMed, EMBASE and the Cochrane Library. Two investigators independently reviewed all identified abstracts and selected articles for full-text review. We included original studies assessing mediators in intra-abdominal sepsis/injury. RESULTS: Among 2437 citations, we selected 182 studies in the scoping review, including 79 preclinical and 103 clinical studies. Serum procalcitonin and C-reactive protein appear to be useful to rule out infection or monitor therapy; however, the diagnostic and prognostic value of mediators for complications/outcomes of sepsis or injury remains to be established. Peritoneal mediator levels are substantially higher than systemic levels after intra-abdominal infection/trauma. Common limitations of current studies included small sample sizes and lack of uniformity in study design and outcome measures. To date, targeted therapies against mediators remain experimental. CONCLUSIONS: Whereas preclinical data suggests mediators play a critical role in intra-abdominal sepsis or injury, there is no consensus on the clinical use of mediators in diagnosing or managing intra-abdominal sepsis or injury. Measurement of peritoneal mediators should be further investigated as a more sensitive determinant of intra-abdominal inflammatory response. High-quality clinical trials are needed to better understand the role of inflammatory mediators.

Active Negative Pressure Peritoneal Therapy After Abbreviated Laparotomy: The Intraperitoneal Vacuum Randomized Controlled Trial.

OBJECTIVE: To determine whether active negative pressure peritoneal therapy with the ABThera temporary abdominal closure device reduces systemic inflammation after abbreviated laparotomy. BACKGROUND: Excessive systemic inflammation after abdominal injury or intra-abdominal sepsis is associated with poor outcomes. METHODS: We conducted a single-center, randomized controlled trial. Forty-five adults with abdominal injury (46.7%) or intra-abdominal sepsis (52.3%) were randomly allocated to the ABThera (n = 23) or Barker's vacuum pack (n = 22). On study days 1, 2, 3, 7, and 28, blood and peritoneal fluid were collected. The primary endpoint was the difference in the plasma concentration of interleukin-6 (IL-6) 24 and 48 hours after temporary abdominal closure application. RESULTS: There was a significantly lower peritoneal fluid drainage from the ABThera at 48 hours after randomization. Despite this, there was no difference in plasma concentration of IL-6 at baseline versus 24 (P = 0.52) or 48 hours (P = 0.82) between the groups. There was also no significant intergroup difference in the plasma concentrations of IL-1ß, -8, -10, or -12 p70 or tumor necrosis factor α between these time points. The cumulative incidence of primary fascial closure at 90 days was similar between groups (hazard ratio, 1.6; 95% confidence interval, 0.82-3.0; P = 0.17). However, 90-day mortality was improved in the ABThera group (hazard ratio, 0.32; 95% confidence interval, 0.11-0.93; P = 0.04). CONCLUSIONS: This trial observed a survival difference between patients randomized to the ABThera versus Barker's vacuum pack that did not seem to be mediated by an improvement in peritoneal fluid drainage, fascial closure rates, or markers of systemic inflammation. TRIAL REGISTRATION: ClinicalTrials.gov identifier NCT01355094.

Does trauma team activation associate with the time to CT scan for those suspected of serious head injuries?

BACKGROUND: Traumatic brain injury (TBI) constitutes the leading cause of posttraumatic mortality. Practically, the major interventions required to treat TBI predicate expedited transfer to CT after excluding other immediately life-threatening conditions. At our center, trauma responses variably consist of either full trauma activation (FTA) including an attending trauma surgeon or a non-trauma team response (NTTR). We sought to explore whether FTAs expedited the time to CT head (TTCTH). METHODS: Retrospective review of augmented demographics of 88 serious head injuries identified from a Regional Trauma Registry within one year at a level I trauma center. The inclusion criteria consisted of a diagnosis of head injury recorded as intubated or GCS < 13; and CT-head scanning after arriving the emergency department. Data was analyzed using STATA. RESULTS: There were 58 FTAs and 30 NTTRs; 86% of FTAs and 17% of NTTRs were intubated prehospital out of 101 charts reviewed in detail; 13 were excluded due to missing data. Although FTAs were more seriously injured (median ISS 29, MAIS head 19, GCS score at scene 6.0), NTTRs were also severely injured (median ISS 25, MAIS head 21, GCS at scene 10) and older (median 54 vs. 26 years). Median TTCTH was double without dedicated FTA (median 50 vs. 26 minutes, p < 0.001), despite similar justifiable delays (53% NTTR, 52% FTA). Without FTA, most delays (69%) were for emergency intubation. TTCTH after securing the airway was longer for NTTR group (median 38 vs. 26 minutes, p =0.0013). Even with no requirements for ED interventions, TTCTH for FTA was less than half versus NTTR (25 vs. 61 minutes, p =0.0013). Multivariate regression analysis indicated age and FTA with an attending surgeon as significant predictors of TTCTH, although the majority of variability in TTCTH was not explained by these two variables (R² = 0.33). CONCLUSION: Full trauma activations involving attending trauma surgeons were quicker at transferring serious head injury patients to CT. Patients with FTA were younger and more seriously injured. Discerning the reasons for delays to CT should be used to refine protocols aimed at minimizing unnecessary delays and enhancing workforce efficiency and clinical outcome.

Help is in your pocket: the potential accuracy of smartphone- and laptop-based remotely guided resuscitative telesonography.

BACKGROUND: Ultrasound (US) examination has many uses in resuscitation, but to use it to its full effectiveness typically requires a trained and proficient user. We sought to use information technology advances to remotely guide US-naive examiners (UNEs) using a portable battery-powered tele-US system mentored using either a smartphone or laptop computer. MATERIALS AND METHODS: A cohort of UNEs (5 tactical emergency medicine technicians, 10 ski-patrollers, and 4 nurses) was guided to perform partial or complete Extended Focused Assessment with Sonography of Trauma (EFAST) examinations on both a healthy volunteer and on a US phantom, while being mentored by a remote examiner who viewed the US images over either an iPhone(®) (Apple, Cupertino, CA) or a laptop computer with an inlaid depiction of the US probe and the "patient," derived from a videocamera mounted on the UNE's head. Examinations were recorded as still images and over-read from a Web site by seven expert reviewers (ERs) (three surgeons, two emergentologists, and two radiologists). Examination goals were to identify lung sliding (LS) documented by color power Doppler (CPD) in the human and to identify intraperitoneal (IP) fluid in the phantom. RESULTS: All UNEs were successfully mentored to easily and clearly identify both LS (19 determinations) and IP fluid (14 determinations), as assessed in real time by the remote mentor. ERs confirmed IP fluid in 95 of 98 determinations (97%), with 100% of ERs perceiving clinical utility for the abdominal Focused Assessment with Sonography of Trauma. Based on single still CPD images, 70% of ERs agreed on the presence or absence of LS. In 16 out of 19 cases, over 70% of the ERs felt the EFAST exam was clinically useful. CONCLUSIONS: UNEs can confidently be guided to obtain critical findings using simple information technology resources, based on the receiving/transmitting device found in most trauma surgeons' pocket or briefcase. Global US mentoring requires only Internet connectivity and initiative.

The feasibility of nurse practitioner-performed, telementored lung telesonography with remote physician guidance - 'a remote virtual mentor'.

BACKGROUND: Point-of-care ultrasound (POC-US) use is increasingly common as equipment costs decrease and availability increases. Despite the utility of POC-US in trained hands, there are many situations wherein patients could benefit from the added safety of POC-US guidance, yet trained users are unavailable. We therefore hypothesized that currently available and economic 'off-the-shelf' technologies could facilitate remote mentoring of a nurse practitioner (NP) to assess for recurrent pneumothoraces (PTXs) after chest tube removal. METHODS: The simple remote telementored ultrasound system consisted of a handheld ultrasound machine, head-mounted video camera, microphone, and software on a laptop computer. The video output of the handheld ultrasound machine and a macroscopic view of the NP's hands were displayed to a remote trauma surgeon mentor. The mentor instructed the NP on probe position and US machine settings and provided real-time guidance and image interpretation via encrypted video conferencing software using an Internet service provider. Thirteen pleural exams after chest tube removal were conducted. RESULTS: Thirteen patients (26 lung fields) were examined. The remote exam was possible in all cases with good connectivity including one trans-Atlantic interpretation. Compared to the subsequent upright chest radiograph, there were 4 true-positive remotely diagnosed PTXs, 2 false-negative diagnoses, and 20 true-negative diagnoses for 66% sensitivity, 100% specificity, and 92% accuracy for remotely guided chest examination. CONCLUSIONS: Remotely guiding a NP to perform thoracic ultrasound examinations after tube thoracostomy removal can be simply and effectively performed over encrypted commercial software using low-cost hardware. As informatics constantly improves, mentored remote examinations may further empower clinical care providers in austere settings.

Efficacy and safety of active negative pressure peritoneal therapy for reducing the systemic inflammatory response after damage control laparotomy (the Intra-peritoneal Vacuum Trial): study protocol for a randomized controlled trial.

BACKGROUND: Damage control laparotomy, or abbreviated initial laparotomy followed by temporary abdominal closure (TAC), intensive care unit resuscitation, and planned re-laparotomy, is frequently used to manage intra-abdominal bleeding and contamination among critically ill or injured adults. Animal data suggest that TAC techniques that employ negative pressure to the peritoneal cavity may reduce the systemic inflammatory response and associated organ injury. The primary objective of this study is to determine if use of a TAC dressing that affords active negative pressure peritoneal therapy, the ABThera Open Abdomen Negative Pressure Therapy System, reduces the extent of the systemic inflammatory response after damage control laparotomy for intra-abdominal sepsis or injury as compared to a commonly used TAC method that provides potentially less efficient peritoneal negative pressure, the Barker's vacuum pack. METHODS/DESIGN: The Intra-peritoneal Vacuum Trial will be a single-center, randomized controlled trial. Adults will be intraoperatively allocated to TAC with either the ABThera or Barker's vacuum pack after the decision has been made by the attending surgeon to perform a damage control laparotomy. The study will use variable block size randomization. On study days 1, 2, 3, 7, and 28, blood will be collected. Whenever possible, peritoneal fluid will also be collected at these time points from the patient's abdomen or TAC device. Luminex technology will be used to quantify the concentrations of 65 mediators relevant to the inflammatory response in peritoneal fluid and plasma. The primary endpoint is the difference in the plasma concentration of the pro-inflammatory cytokine IL-6 at 24 and 48 h after TAC dressing application. Secondary endpoints include the differential effects of these dressings on the systemic concentration of other pro-inflammatory cytokines, collective peritoneal and systemic inflammatory mediator profiles, postoperative fluid balance, intra-abdominal pressure, and several patient-important outcomes, including organ dysfunction measures and mortality. DISCUSSION: Results from this study will improve understanding of the effect of active negative pressure peritoneal therapy after damage control laparotomy on the inflammatory response. It will also gather necessary pilot information needed to inform design of a multicenter trial comparing clinical outcomes among patients randomized to TAC with the ABThera versus Barker's vacuum pack. TRIAL REGISTRATION: ClinicalTrials.gov identifier http://www.clicaltrials.gov/ct2/show/NCT01355094.

Enabling the mission through trans-atlantic remote mentored musculoskeletal ultrasound: case report of a portable hand-carried tele-ultrasound system for medical relief missions.

Modern medical practice has become extremely dependent upon diagnostic imaging technologies to confirm the results of clinical examination and to guide the response to therapies. Of the various diagnostic imaging techniques, ultrasound is the most portable modality and one that is repeatable, dynamic, relatively cheap, and safe as long as the imaging provided is accurately interpreted. It is, however, the most user-dependent, a characteristic that has prompted the development of remote guidance techniques, wherein remote experts guide distant users through the use of information technologies. Medical mission work often brings specialist physicians to less developed locations, where they wish to provide the highest levels of care but are often bereft of diagnostic imaging resources on which they depend. Furthermore, if these personnel become ill or injured, their own care received may not be to the standard they have left at home. We herein report the utilization of a compact hand-carried remote tele-ultrasound system that allowed real-time diagnosis and follow-up of an acutely torn adductor muscle by a team of ultrasonographers, surgeons, and physicians. The patient was one of the mission surgeons who was guided to self-image. The virtual network of supporting experts was located across North America, whereas the patient was in Lome, Togo, West Africa. The system consisted of a hand-carried ultrasound, the output of which was digitized and streamed to the experts within standard voice-over-Internet-protocol software with an embedded simultaneous videocamera image of the ultrasonographer's hands using a customized graphical user interface. The practical concept of a virtual tele-ultrasound support network was illustrated through the clinical guidance of multiple physicians, including National Aeronautics and Space Administration Medical Operations remote guiders, Olympic team-associated surgeons, and ultrasound-focused emergentologists.

Occult pneumothoraces in critical care: a prospective multicenter randomized controlled trial of pleural drainage for mechanically ventilated trauma patients with occult pneumothoraces.

BACKGROUND: Patients with an occult pneumothoraces (OPTXs) may be at risk of tension pneumothoraces (TPTXs) without drainage or pleural drainage complications if treated. METHODS: Adults with traumatic OPTXs and requiring positive-pressure ventilation (PPV) were randomized to pleural drainage or observation (one side only enrolled if bilateral). All subsequent care and method of pleural drainage was per attending physician discretion. The primary outcome was a composite of respiratory distress (RD) (need for urgent pleural drainage, acute/sustained increases in O2 requirements, ventilator dysynchrony, and/or charted respiratory events). RESULTS: Ninety severely injured patients (mean [SD], Injury Severity Score [ISS], 33 [11]) were studied at four centers: Calgary (55), Toronto (27), Quebec (6), and Sherbrooke (3). Forty were randomized to tube thoracostomy, and 50 were randomized to observation. The risk of RD was similar between the observation and tube thoracostomy groups (relative risk, 0.71; 95% confidence interval, 0.40-1.27). There was no difference in mortality or intensive care unit (ICU), ventilator, or hospital days between groups. In those observed, 20% required subsequent pleural drainage (40% PTX progression, 60% pleural fluid, and 20% other). One observed patient (2%) undergoing PPV at enrollment had a TPTX, which was treated with urgent tube thoracostomy without sequelae. Drainage complications occurred in 15% of those randomized to drainage, while suboptimal tube thoracostomy position occurred in an additional 15%. There were three times (24% vs. 8%) more failures and more RDs (p = 0.01) among those observed with OPTXs requiring sustained PPV versus just for an operation, which increases threefold after a week in the ICU (p = 0.07). CONCLUSION: Our results suggest that OPTXs may be safely observed in hemodynamically stable patients undergoing PPV just for an operation, although one third of those requiring a week or more of ICU care received drainage, and TPTXs still occur. Complications of pleural drainage remain unacceptably high, and future work should attempt to delineate specific factors among those observed that warrant prophylactic drainage. LEVEL OF EVIDENCE: Therapeutic study, level III.

Potential Use of Remote Telesonography as a Transformational Technology in Underresourced and/or Remote Settings.

Mortality and morbidity from traumatic injury are twofold higher in rural compared to urban areas. Furthermore, the greater the distance a patient resides from an organized trauma system, the greater the likelihood of an adverse outcome. Delay in timely diagnosis and treatment contributes to this penalty, regardless of whether the inherent barriers are geographic, cultural, or socioeconomic. Since ultrasound is noninvasive, cost-effective, and portable, it is becoming increasingly useful for remote/underresourced (R/UR) settings to avoid lengthy patient travel to relatively inaccessible medical centers. Ultrasonography is a user-dependent, technical skill, and many, if not most, front-line care providers will not have this advanced training. This is particularly true if care is being provided by out-of-hospital, "nontraditional" providers. The human exploration of space has forced the utilization of information technology (IT) to allow remote experts to guide distant untrained care providers in point-of-care ultrasound to diagnose and manage both acute and chronic illness or injuries. This paradigm potentially brings advanced diagnostic imaging to any medical interaction in a setting with internet connectivity. This paper summarizes the current literature surrounding the development of teleultrasound as a transformational technology and its application to underresourced settings.

A molecular complex of bovine milk protein and oleic acid selectively kills cancer cells in vitro and inhibits tumour growth in an orthotopic rat bladder tumour model.

UNLABELLED: WHAT'S KNOWN ON THE SUBJECT? AND WHAT DOES THE STUDY ADD?: Novel intravesical therapies are needed for superficial bladder cancer that reduce the risk of infection associated with Bacillus Calmette-Guérin (BCG) and further destabilization of the urothelium associated with cytotoxic chemotherapy. Experimental therapies to date have included photodynamic therapy, oncolytic viruses, gene therapy (antisense oligonucleotides and silencing RNA), cytokine therapy, death receptor agonists (tumour-necrosis-factor-related apoptosis-inducing ligand and anti-DR5 monoclonal antibody), naturally occurring substances (curcumin and deguelin) and human α-lactalbumin made lethal to tumour cells (HAMLET). HAMLET, a natural occurring product in milk, induces apoptosis in urothelial cancer cells but has limitations in clinical application because of its human source. A previous study in patients with bladder cancer has demonstrated that intravesical HAMLET (daily for 5 days before tumour resection) caused selective apoptotic tumour cell death. BAMLET, the bovine equivalent of HAMLET, is a complex of bovine α-lactalbumin and oleic acid (bLAC) that has been shown in vitro to accumulate in the endolysosomal compartment of tumour cells and induce leakage of lysosomal cathepsins into the cytosol followed by activation of the pro-apoptotic protein Bax. This is the first in vivo study to show that BAMLET (bLAC) induces apoptosis in urothelial cancer cells and controls the growth of high risk urothelial cancer in a syngeneic rat orthotopic model. This same bladder cancer model system has been used to test other novel therapies, including BCG, and therefore provides a relative comparison of its effectiveness with other intravesical therapies. OBJECTIVE: To investigate the efficacy of a complex of bovine α-lactalbumin and oleic acid (bLAC) to kill urothelial cancer cells in vitro and inhibit tumour growth and progression in a high risk bladder tumour model. MATERIALS AND METHODS: The cytotoxicity of bLAC to a large panel of urothelial cell cancer (UCC) cells was tested by the MTT (3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide) assay, using bLA, the folded α-lactalbumin without oleic acid, as a control. The mechanism of bLAC-inducing cell death was evaluated by annexin V staining, TUNEL (terminal deoxynucleotidyl transferase mediated nick end labelling) assay and sub-G1 DNA analysis. The selective bLAC cytotoxicity was examined using multicellular spheroids consisting of UCC and non-transformed fibroblasts. Rats bearing orthotopic tumour received intravesical instillations (twice weekly, for 3 weeks) of bLAC, bLA, BCG or saline, starting 6 days after UCC (AY-27) cell inoculation. Animals were monitored for survival, toxicity and tumour growth control. RESULTS: A dose-dependent bLAC-inducing apoptotic-like cell death was shown in UCC cells tested, including cells refractory to classic apoptosis-inducing agents, whereas bLA showed little cytotoxicity. bLAC selectively destroyed cancer cells in spheroids. Intravesical bLAC therapy demonstrated marked reduction in tumour growth/progression and significantly prolonged animal survival vs saline instillations (P = 0.004, log-rank test) and showed comparable efficacy with BCG (standard) therapy. CONCLUSION: The findings identify bLAC as a new candidate for UCC therapy and suggests that topical administration of bLAC alone or with BCG to prevent progression of bladder cancer warrants further exploration.

Antitumor Efficacy of Intravesical BCG, Gemcitabine, Interferon-α and Interleukin-2 as Mono- or Combination-Therapy for Bladder Cancer in an Orthotopic Tumor Model.

OBJECTIVE: To reduce adverse effects and improve efficacy of intravesical BCG for bladder cancer, alternative treatment options were investigated in an orthotopic rat tumor model. METHODS: Superficial bladder cancer was established in syngeneic female rat bladders by instillation of AY-27 cells. Animals were randomly assigned to treatment groups including dose escalation of intravesical BCG with or without interferon-α (IFN-α) or interleukin-2 (IL-2); or graded doses of gemcitabine alone; or BCG plus gemcitabine. Treatments were given twice weekly for 3 weeks. Rats in control groups received saline instillations. Treatment response was monitored by animals' well-being, survival days, tumor growth inhibition, and histological examination at necropsy. RESULTS: Rats receiving monotherapy with intravesical BCG, gemcitabine, or IFN-α, attained significantly better survival and tumor reduction compared with control (P = 0.002; 0.001; 0.002, respectively, Log-rank Test). A dose-dependent treatment response was observed in animals with established bladder tumor receiving escalated BCG instillations. Only high-dose BCG significantly improved animal survival. Although high-dose BCG plus gemcitabine or IFN-α did not increase benefit over monotherapies, low-dose BCG plus IL-2 did show improved efficacy (P = 0.01). CONCLUSION: Intravesical monotherapies with gemcitabine and IFN-α were as effective as BCG for treatment of early non-muscle-invasive urothelial bladder cancer in this immune competent rat model. Combining these agents with high-dose BCG did not further increase efficacy. However, combining low-dose BCG with IL-2 enhanced BCG effectiveness.

Lipophilic photosensitizer administration via the prostate arteries for photodynamic therapy of the canine prostate.

BACKGROUND: Current limitations of interstitial photodynamic therapy (PDT) for treatment of prostate cancer include low drug selectivity after intravenous (i.v.) administration and incomplete ablation of glandular tissue. To overcome these limitations, intra-arterial (i.a.) injection of a photosensitizer was tested in a canine model. METHODS: A lipophilic photosensitizer, SL052 formulated in liposomes or dissolved in dimethyl sulphoxide (DMSO), was injected into male dogs as an intravenous injection or intra-arterially via the prostate arteries. Optical fibers were inserted into the prostate 3h after i.v. or immediately following i.a. drug delivery. Laser light was delivered through the fibers in cycles controlled by a computer-driven switch. Drug concentration (fluorescence) and light transmission in prostate tissue were monitored during the course of PDT. Side effect profile and completeness of prostate gland ablation were the primary parameters compared among treatment groups. Control animals received drug-only or light-only treatment. RESULTS AND CONCLUSION: Thirteen dogs were treated by PDT mediated by i.a. injection of SL052 dissolved in DMSO and attained either complete ablation of prostatic glandular tissue or significant reduction of prostate volume compared with that of pre-PDT (p<0.0001). When compared to i.v. administration the i.a. route resulted in more complete photo-ablation. Associated side effect included acute urinary retention which resolved overtime. No incontinence was observed. With careful tailoring of PDT drug and light doses, interstitial PDT with i.a. injection of SL052-DMSO has the potential to provide effective treatment for prostate disease.

Response of bladder carcinoma cells to TRAIL and antisense oligonucleotide, Bcl-2 or clusterin treatments.

PURPOSE: Bladder transitional cell carcinoma is the second most common urological malignancy, of which 80% are superficial disease limited to the bladder. Superficial bladder transitional cell carcinoma has a high propensity for recurrence and progression after initial resection, necessitating adjuvant intravesical therapy. TRAIL (tumor necrosis factor-related apoptosis inducing ligand) can selectively induce apoptosis in most tumor cells while sparing normal cells. TRAIL drives not only the death receptor pathway, but also the mitochondrial pathway through Bid. Due to the anti-apoptotic functions of Bcl-2 and clusterin on the mitochondrial apoptotic pathway the effects of down-regulating these proteins were examined in partially TRAIL resistant bladder transitional cell carcinoma cell lines. MATERIALS AND METHODS: Antisense oligonucleotides targeting Bcl-2 and clusterin were used alone or combined with TRAIL and cytotoxicity was examined by the MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolim bromide) proliferation assay. Apoptotic pathway signals were detected by sodium dodecyl sulfate-polyacrylamide gel electrophoresis/Western blotting after the various combination treatments. All statistical tests were 2 sided. RESULTS: Although no direct correlation between TRAIL sensitivity and the relative expression levels of Bcl-2 and clusterin was found in the bladder transitional cell carcinoma cell lines examined, antisense oligonucleotide mediated the down-regulation of Bcl-2 and clusterin, increasing the sensitivity of the partially resistant cells to TRAIL. This was mediated through increased apoptotic signaling of the mitochondrial pathway, as evident by the increased activation of caspase-9 and 3, and cleaved DFF45. There was no benefit of combined antisense oligonucleotide therapy. CONCLUSIONS: This study provides proof of principle that TRAIL combined with antisense oligonucleotide-Bcl-2 may have potential as a novel future treatment strategy for bladder transitional cell carcinoma.

Photodynamic therapy of the canine prostate: intra-arterial drug delivery.

PURPOSE: Interstitial photodynamic therapy (PDT) selectively destroys tissue targeted with a photosensitizer and then exposed to light of a specific wavelength. We report a novel delivery method--intra-arterial drug delivery for PDT of the prostate--in a canine model. METHODS: To evaluate drug distribution, the prostatovesical artery was selectively cannulated and photosensitizers alone or in conjunction with 99m-technetium-labeled macro-aggregated albumin ((99m)Tc-MAA) were injected via a 3 Fr microcatheter in 8 animals. One dog was followed for 3 months to determine tolerance and toxicity. The remaining animals were euthanized and imaged with whole-body single photon emission CT and gamma counting for radioactivity distribution. Photosensitizer distribution was further analyzed by fluorescence confocal microscopy and tissue chemical extraction. To evaluate PDT, the photosensitizer QLT0074 was infused in 3 animals followed by interstitial illumination with 690 nm laser light. RESULTS: Intra-arterial infusion selectively delivered drugs to the prostate, with both radioactivity and photosensitizer levels significantly higher (up to 18 times) than in the surrounding organs (i.e., rectum). With unilateral injection of (99m)Tc-MAA, only the injected half of the prostate showed activity whereas bilateral administration resulted in drug delivery to the entire prostate. PDT resulted in comprehensive damage to the prostate without severe complications or systemic toxicity. CONCLUSION: Injection of radiolabeled MAA into the prostatovesical artery results in distribution within the prostate with negligible amounts reaching the adjacent organs. PDT also demonstrates selective damage to the prostate, which warrants clinical application in targeted prostate therapies.

Fractionated versus standard continuous light delivery in interstitial photodynamic therapy of dunning prostate carcinomas.

PURPOSE: The study aims to compare the standard/continuous light delivery with fractionated light delivery for interstitial photodynamic therapy (PDT) of prostate cancer. EXPERIMENTAL DESIGN: Dunning R3327 prostate tumor models were established in male syngeneic rats. When tumors reached approximately 3,000 mm3, animals were randomized to various treatment groups. Three hours after QLT0074 injection, tumors were illuminated by 690-nm light delivered by a computer-controlled switch, which sequentially directed light to one of the seven optical fibers in cycles. For comparison, tumors were treated with continuous illumination. Tumors treated with light-only served as control. Dynamic contrast-enhanced magnetic resonance imaging was used to monitor tumor perfusion changes before and after PDT. RESULTS: Tumor response (animal survival) to PDT with fractionated light delivery was PDT dose dependent in both tumor models. Rats bearing anaplastic tumor treated by fractionated light (PDT dose: 1.5 mg/kg QLT0074, 900 J light) had a median survival of 51 days with 25% tumor cures compared with that of 26 days with no tumor cure by continuous illumination (P = 0.015) and 14 days by light-only (P = 0.0001). Rats bearing well-differentiated tumor treated by fractionated light had a median survival of 82 days compared with 65 days by continuous illumination (P = 0.001) and 37 days by light-only. PDT with fractionated light generated a perfusion reduction of 80% compared with 52% for continuous illumination in well-differentiated tumors. CONCLUSIONS: Fractionated light delivery is more effective than continuous light delivery in PDT of prostate cancer (solid tumors). These results warrant further investigation in clinical trials.