Sonic hedgehog suppresses penile remodeling after cavernous nerve injury and sustains long-term normal penis morphology.

BACKGROUND: Cavernous nerve (CN) injury, which occurs in prostatectomy and diabetic cases, initiates penile remodeling, including smooth muscle apoptosis and increased collagen in the corpora cavernosa, which are underlying causes of erectile dysfunction. Sonic hedgehog (SHH) is a critical regulator of penile smooth muscle, and SHH treatment suppresses corpora cavernosa remodeling that occurs with CN injury. AIM: We examine if SHH treatment by peptide amphiphile (PA) in the first week after CN injury is sufficient to prevent long-term penis remodeling and if apoptosis inhibitors also suppress penile remodeling. METHODS: Bilateral CN crush was performed on adult Sprague-Dawley rats (P115-120) that underwent 1 of 3 treatments with novel extended-release nanofiber PA hydrogels for delivery: SHH protein (n = 10), mouse serum albumin protein (control, n = 7), or caspase 3 inhibitor (AC-DEVD-CHO, n = 10). Rats were sacrificed after 18 to 24 days. Additional rats underwent CN injury (n = 6) or CN injury and SHH PA treatment for 2 and 4 days (n = 8) and included sham controls (n = 3) and nonsurgery controls (n = 3). OUTCOMES: Trichrome stain, hydroxyproline assay, and Western analysis for α-actin (smooth muscle) and GAPDH were performed to examine smooth muscle retention and collagen abundance. RESULTS: Smooth muscle decreased with CN injury. Corpora cavernosa showed increased smooth muscle at 2, 4, and 24 days after CN injury with SHH PA treatment in comparison with mouse serum albumin treatment among CN-injured controls. Caspase 3-inhibited penis demonstrated little smooth muscle preservation. Collagen was decreased 23% with SHH PA treatment (P < .001) at 18 to 24 days after CN injury. Collagen was unchanged with caspase 3 inhibitor treatment (P > .99). CLINICAL TRANSLATION: It is important to know that treatments given at the time of CN injury have a sustained effect on preserving penile architecture and thus erectile function, making them valuable for clinical translation. STRENGTHS AND LIMITATIONS: SHH PA treatment preserves penile smooth muscle after CN injury. Time points past 24 days were not examined, although penile remodeling takes place acutely after CN injury. Measurement of erectile function was not examined. CONCLUSIONS: SHH treatment by PA in the first week after CN injury is sufficient to suppress penile remodeling and to preserve penile smooth muscle over time, which is critical to prevent development of erectile dysfunction. There is a difference in the corpora cavernosa smooth muscle from proximal to distal in the penis of the Sprague-Dawley rat model. It is critical when examining therapy efficacy to ensure that comparable regions of the penis are analyzed. STATEMENT OF SIGNIFICANCE: In this study, we examine if suppression of apoptosis in penile smooth muscle in the first week after cavernous nerve injury is sufficient to preserve smooth muscle long-term.

SHH regulates penile morphology and smooth muscle through a mechanism involving BMP4 and GREM1.

BACKGROUND: The cavernous nerve (CN) is frequently damaged in prostatectomy and diabetic patients with erectile dysfunction (ED), initiating changes in penile morphology including an acute and intense phase of apoptosis in penile smooth muscle and increased collagen, which alter penile architecture and make corpora cavernosa smooth muscle less able to relax in response to neurotransmitters, resulting in ED. AIM: Sonic hedgehog (SHH) is a critical regulator of penile smooth muscle, and SHH treatment suppresses penile remodeling after CN injury through an unknown mechanism; we examine if part of the mechanism of how SHH preserves smooth muscle after CN injury involves bone morphogenetic protein 4 (BMP4) and gremlin1 (GREM1). METHODS: Primary cultures of smooth muscle cells were established from prostatectomy, diabetic, hypertension and Peyronie's (control) (N = 18) patients. Cultures were characterized by ACTA2, CD31, P4HB, and nNOS immunohistochemical analysis. Patient smooth muscle cell growth was quantified in response to BMP4 and GREM1 treatment. Adult Sprague Dawley rats underwent 1 of 3 surgeries: (1) uninjured or CN-injured rats were treated with BMP4, GREM1, or mouse serum albumin (control) proteins via Affi-Gel beads (N = 16) or peptide amphiphile (PA) (N = 26) for 3 and 14 days, and trichrome stain was performed; (2) rats underwent sham (N = 3), CN injury (N = 9), or CN injury and SHH PA treatment for 1, 2, and 4 days (N = 9). OUTCOMES: Western analysis for BMP4 and GREM1 was performed; (3) rats were treated with 5E1 SHH inhibitor (N = 6) or IgG (control; N = 6) for 2 and 4 days, and BMP4 and GREM1 localization was examined. Statistics were performed by analysis of variance with Scheffé's post hoc test. RESULTS: BMP4 increased patient smooth muscle cell growth, and GREM1 decreased growth. In rats, BMP4 treatment via Affi-Gel beads and PA increased smooth muscle at 3 and 14 days of treatment. GREM1 treatment caused increased collagen and smooth muscle at 3 days, which switched to primarily collagen at 14 days. CN injury increased BMP4 and GREM1, while SHH PA altered Western band size, suggesting alternative cleavage and range of BMP4 and GREM1 signaling. SHH inhibition in rats increased BMP4 and GREM1 in fibroblasts. CLINICAL IMPLICATIONS: Understanding how SHH PA preserves and regenerates penile morphology after CN injury will aid development of ED therapies. STRENGTHS AND LIMITATIONS: SHH treatment alters BMP4 and GREM1 localization and range of signaling, which can affect penile morphology. CONCLUSION: Part of the mechanism of how SHH regulates corpora cavernosa smooth muscle involves BMP4 and GREM1.

BMP4 and GREM1 are targets of SHH signaling and downstream regulators of collagen in the penis.

BACKGROUND: Cavernous nerve (CN) injury, caused by prostatectomy and diabetes, initiates a remodeling process (smooth muscle apoptosis and increased collagen) in the corpora cavernosa of the penis of patients and animal models that is an underlying cause of erectile dysfunction (ED), and the Sonic hedgehog (SHH) pathway plays an essential role in the response of the penis to denervation, as collagen increases with SHH inhibition and decreases with SHH treatment. AIM: We examined if part of the mechanism of how SHH prevents penile remodeling and increased collagen with CN injury involves bone morphogenetic protein 4 (BMP4) and gremlin1 (GREM1) and examined the relationship between SHH, BMP4, GREM1, and collagen in penis of ED patients and rat models of CN injury, SHH inhibition, and SHH, BMP4, and GREM1 treatment. METHODS: Corpora cavernosa of Peyronie's disease (control), prostatectomy, and diabetic ED patients were obtained (N = 30). Adult Sprague Dawley rats (n = 90) underwent (1) CN crush (1-7 days) or sham surgery; (2) CN injury and BMP4, GREM1, or mouse serum albumin (control) treatment via Affi-Gel beads or peptide amphiphile (PA) for 14 days; (3) 5E1 SHH inhibitor, IgG, or phosphate-buffered saline (control) treatment for 2 to 4 days; or (4) CN crush with mouse serum albumin or SHH for 9 days. OUTCOMES: Immunohistochemical and Western analysis for BMP4 and GREM1, and collagen analysis by hydroxyproline and trichrome stain were performed. RESULTS: BMP4 and GREM1 proteins were identified in corpora cavernosa smooth muscle of prostatectomy, diabetic, and Peyronie's patients, and in rat smooth muscle, sympathetic nerve fibers, perineurium, blood vessels, and urethra. Collagen decreased 25.4% in rats with CN injury and BMP4 treatment (P = .02) and increased 61.3% with CN injury and GREM1 treatment (P = .005). Trichrome stain showed increased collagen in rats treated with GREM1. Western analysis identified increased BMP4 and GREM1 in corpora cavernosa of prostatectomy and diabetic patients, and after CN injury (1-2 days) in our rat model. Localization of BMP4 and GREM1 changed with SHH inhibition. SHH treatment increased the monomer form of BMP4 and GREM1, altering their range of signaling. CLINICAL IMPLICATIONS: A better understanding of penile remodeling and how fibrosis occurs with loss of innervation is essential for development of novel ED therapies. STRENGTHS AND LIMITATIONS: The relationship between SHH, BMP4, GREM1, and collagen is complex in the penis. CONCLUSION: BMP4 and GREM1 are downstream targets of SHH that impact collagen and may be useful in collaboration with SHH to prevent penile remodeling and ED.

Motor vehicle collision (MVC) emergency department (ED) visits and hospitalisations in Ontario during the COVID-19 pandemic.

BACKGROUND: The COVID-19 pandemic policy response dramatically changed local transportation patterns. This project investigated the impact of COVID-19 policies on motor vehicle collision (MVC)-related emergency department (ED) visits and hospitalisations in Ontario. METHODS: Data were collected on MVC-related ED visits and hospitalisations in Ontario between March 2016 and December 2022. Using an interrupted time series design, negative binomial regression models were fitted to the pre-pandemic data, including monthly indicator variables for seasonality and accounting for autocorrelation. Extrapolations simulated expected outcome trajectories during the pandemic, which were compared with actual observed outcome counts using the overall per cent change and mean monthly difference. Data were modelled separately for vehicle occupants, pedestrians and cyclists (MVC and non-MVC injuries). RESULTS: There was a 31.5% decrease in observed ED visits (95% CI -35.4 to -27.3) and a 6.0% decrease in hospitalisations (95% CI -13.2 to 1.6) among vehicle occupants, relative to expected counts during the pandemic. Results were similar for pedestrians. Among cyclist MVCs, there was an increase in ED visits (12.8%, 95% CI -8.2 to 39.4) and hospitalisations (46.0%, 95% CI 11.6 to 93.6). Among non-MVC cyclists, there was also an increase in ED visits (47.0%, 95% CI 12.5 to 86.8) and hospitalisations (50.1%, 95% CI 8.2 to 101.2). CONCLUSIONS: We observed fewer vehicle occupant and pedestrian collision injuries than expected during the pandemic. By contrast, we observed more cycling injuries than expected, especially in cycling injuries not involving motor vehicles. These observations may be attributable to changes in transportation patterns during the pandemic and increased uptake of recreational cycling.

Supporting Ontario public health units to address adverse childhood experiences in pandemic recovery planning: A priority-setting exercise.

BACKGROUND: Adverse childhood experiences (ACEs) are potentially traumatic exposures experienced during childhood, for example, neglect. There is growing evidence that the coronavirus disease 2019 (COVID-19) pandemic and related socioeconomic conditions contributed to an increased risk of ACEs. As public health programs/services are re-evaluated and restored following the state of emergency, it is important to plan using an ACEs-informed lens. The aim of this study was to identify and prioritize initiatives or activities that Public Health Ontario (PHO) could undertake to support Ontario public health units' work towards ACEs-informed pandemic recovery plans. METHODS: The Child Health and Nutrition Research Initiative method was adapted to conduct a priority-setting exercise (May-October 2022). Two online surveys were administered with members of the Healthy Growth and Development (HGD) Evidence Network, comprised of public health unit staff working in child and family health/HGD from Ontario's 34 public health units. In the first survey, participants were asked to propose activities or initiatives that PHO could undertake to support Ontario public health units' work towards ACEs-informed planning. In the second survey, participants were asked to score the final list of options against pre-determined prioritization criteria (for example, relevance). Responses were numerically coded and used to calculate prioritization scores, which were used to rank the options. RESULTS: In all, 76% of public health units (n = 26) responded to the first survey to identify options. The 168 proposed ideas were consolidated into a final list of 13 options, which fall under PHO's scientific and technical support mandate areas (data and surveillance, evidence synthesis, collaboration and networking, knowledge exchange and research). A total of 79% of public health units (n = 27) responded to the follow-up survey to prioritize options. Prioritization scores ranged from 76.4% to 88.6%. The top-ranked option was the establishment of a new provincial ACEs community of practice. CONCLUSIONS: Over three quarters of public health units contributed to identifying and ranking 13 options for PHO to support public health units in considering and addressing ACEs through pandemic recovery planning. In consultation with the ACEs and Resilience Community of Practice, recently formed on the basis of this exercise, PHO will continue to use the ranked list of options to inform work-planning activities/priorities.

Evaluation of knowledge-based planning models for male pelvic CBCT-based online adaptive radiotherapy on conventional linear accelerators.

PURPOSE: To assess the practicality of employing a commercial knowledge-based planning tool (RapidPlan) to generate adapted intact prostate and prostate bed volumetric modulated arc therapy (VMAT) plans on iterative cone-beam computed tomography (iCBCT) datasets. METHODS AND MATERIALS: Intact prostate and prostate bed RapidPlan models were trained utilizing planning data from 50 and 44 clinical cases, respectively. To ensure that refined models were capable of producing adequate clinical plans with a single optimization, models were tested with 50 clinical planning CT datasets by comparing dose-volume histogram (DVH) and plan quality metric (PQM) values between clinical and RapidPlan-generated plans. The RapidPlan tool was then used to retrospectively generate adapted VMAT plans on daily iCBCT images for 20 intact prostate and 15 prostate bed cases. As before, DVH and PQM metrics were utilized to dosimetrically compare scheduled (iCBCT Verify) and adapted (iCBCT RapidPlan) plans. Timing data was collected to further evaluate the feasibility of integrating this approach within an online adaptive radiotherapy workflow. RESULTS: Model testing results confirmed the models were capable of producing VMAT plans within a single optimization that were overall improved upon or dosimetrically comparable to original clinical plans. Direct application of RapidPlan on iCBCT datasets produced satisfactory intact prostate and prostate bed plans with generally improved target volume coverage/conformality and rectal sparing relative to iCBCT Verify plans as indicated by DVH values, though bladder metrics were marginally increased on average. Average PQM values for iCBCT RapidPlans were significantly improved compared to iCBCT Verify plans. The average time required [in mm:ss] to generate adapted plans was 06:09 ± 02:06 (intact) and 07:12 ± 01:04 (bed). CONCLUSION: This study demonstrated the feasibility of leveraging RapidPlan to expeditiously generate adapted VMAT intact prostate and prostate bed plans on iCBCT datasets. In general, adapted plans were dosimetrically improved relative to scheduled plans, emphasizing the practicality of the proposed approach.

Monitoring Food Affordability: Reliability and Validity of an Online Nutritious Food Basket.

Purpose: This study aimed to assess the reliability and validity of an online approach to monitoring food affordability in Ontario using the updated Ontario Nutritious Food Basket (ONFB).Methods: The ONFB was priced online in 12 large multi-chain grocery stores to test intra-/inter-rater reliability using percent agreement and intra-class correlations (ICCs). Then, the ONFB was priced in-store and online in 28 stores to estimate food price differences using paired t-tests and Pearson's correlation for all (n =1708) and matched items (same product/brand and purchase unit) (n = 1134).Results: Intra-/inter-rater agreement was high (95.4%/81.6%; ICC = 0.972, F = 69.9, p < 0.001). On average, in-store prices were less than $0.02 lower than online prices. There were no significant differences between mean in-store and online prices for all items (t = 0.504 p = 0.614). The mean price was almost perfectly correlated between in-store and online (fully matched: R = 0.993 p < 0.001; all items: R = 0.967 p < 0.001). Online monthly ONFB estimates for a family of four were strongly correlated (R = 0.937 p < 0.001) with estimates calculated using in-store data.Conclusions: Online pricing is a reliable and valid approach to food costing in Ontario that contributes to modernizing the monitoring of food affordability in Canada and abroad.

Effect of Material and Pad Abrasion on Shear Bond Strength of 3D-Printed Orthodontic Brackets.

OBJECTIVE: To investigate the effect of printing material and air abrasion of bracket pads on the shear bond strength of 3D-printed plastic orthodontic brackets when bonded to the enamel of extracted human teeth. MATERIALS AND METHODS: Premolar brackets were 3D-printed using the design of a commercially available plastic bracket in two biocompatible resins: Dental LT Resin and Dental SG Resin (n = 40/material). 3D-printed brackets and commercially manufactured plastic brackets were divided into two groups (n = 20/group), one of which was air abraded. All brackets were bonded to extracted human premolars, and shear bond strength tests were performed. The failure types of each sample were classified using a 5-category modified adhesive remnant index (ARI) scoring system. RESULTS: Bracket material and bracket pad surface treatment presented statistically significant effects for shear bond strengths, and a significant interaction effect between bracket material and bracket pad surface treatment was observed. The non-air abraded (NAA) SG group (8.87 ± 0.64 MPa) had a statistically significantly lower shear bond strength than the air abraded (AA) SG group (12.09 ± 1.23 MPa). In the manufactured brackets and LT Resin groups, the NAA and AA groups were not statistically significantly different within each resin. A significant effect of bracket material and bracket pad surface treatment on ARI score was observed, but no significant interaction effect between bracket material and pad treatment was found. CONCLUSION: 3D-printed orthodontic brackets presented clinically sufficient shear bond strengths both with and without AA prior to bonding. The effect of bracket pad AA on shear bond strength depends on the bracket material.

Jejunal Inflammation in Crohn's Disease: Comparison between Diffusion Weighted Magnetic Resonance Imaging and Video Capsule Endoscopy.

BACKGROUND: Jejunal disease is associated with worse prognosis in Crohn's disease. The added value of diffusion weighted imaging for evaluating jejunal inflammation related to Crohn's Disease is scarce. OBJECTIVES: To compare diffusion weighted imaging, video capsule endoscopy, and inflammatory biomarkers in the assessment of Crohn's disease involving the jejunum. METHODS: Crohn's disease patients in clinical remission were prospectively recruited and underwent magnetic resonance (MR)-enterography and video capsule endoscopy. C-reactive protein and fecal-calprotectin levels were obtained. MR-enterography images were evaluated for restricted diffusion, and apparent diffusion coefficient values were measured. The video capsule endoscopy-based Lewis score was calculated. Associations between diffusion weighted imaging, apparent diffusion coefficient, Lewis score, and inflammatory biomarkers were evaluated. RESULTS: The study included 51 patients, and 27/51 (52.9%) with video capsule endoscopies showed jejunal mucosal inflammation. Sensitivity and specificity of restricted diffusion for video capsule endoscopy mucosal inflammation were 59.3% and 37.5% for the first reader, and 66.7% and 37.5% for the second reader, respectively. Diffusion weighted imaging was not statistically associated with jejunal video capsule endoscopy inflammation (P = 0.813). CONCLUSIONS: Diffusion weighted imaging was not an effective test for evaluation of jejunal inflammation as seen by video capsule endoscopy in patients with quiescent Crohn's disease.

Sonic Hedgehog Signaling in Primary Culture of Human Corpora Cavernosal Tissue From Prostatectomy, Diabetic, and Peyronie's Patients.

BACKGROUND: Cavernous nerve (CN) injury causes penile remodeling, including smooth muscle apoptosis and increased collagen, which results in erectile dysfunction (ED), and prevention of this remodeling is critical for novel ED therapy development. AIM: We developed 2 peptide amphiphile (PA) hydrogel delivery vehicles for Sonic hedgehog (SHH) protein to the penis and CN, which effectively suppress penile distrophic remodeling (apoptosis and fibrosis), in vivo in a rat CN injury model, and the aim of this study is to determine if SHH PA can be used to regenerate human corpora cavernosal smooth muscle deriving from multiple ED origins. METHODS: Corpora cavernosal tissue was obtained from prostatectomy, diabetic, hypertension, cardiovascular disease and Peyronie's (control) patients (n = 21). Primary cultures (n = 21) were established, and corpora cavernosal cells were treated with SHH protein, MSA (control), 5E1 SHH inhibitor, and PBS (control). Growth was quantified by counting the number of cells at 3-4 days. Statistics were performed by ANOVA with Scheffe's post hoc test. Concentration of SHH protein for maximal growth was optimized, and a more active SHH protein examined. OUTCOMES: Cultures were characterized by immunohistochemical analysis with ACTA2, CD31, nNOS and P4HB, and smooth muscle was quantified in comparison to DAPI. RESULTS: Cultures established were >97% smooth muscle. SHH protein increased growth of smooth muscle cells from prostatectomy, diabetic, and Peyronie's patients in a similar manner (49%-51%), and SHH inhibition decreased growth (20%-33%). There was no difference in growth using 25 ug and 10 ug SHH protein, suggesting a threshold concentration of SHH protein above which smooth muscle growth is enhanced. A more active lipid modified SHH peptide further enhanced growth (15%), indicating a more robust growth response. SHH increased growth in smooth muscle cells from hypertension (37%) and cardiovascular disease (32%) patients. SHH protein increased growth under normal and high glucose conditions, suggesting that high glucose conditions that may be present in under controlled diabetic patients would not detract from SHH regenerative capacity. CLINICAL IMPLICATIONS: SHH PA would be beneficial to enhance smooth muscle regeneration in patients with ED of multiple etiologies. STRENGTHS AND LIMITATIONS: Understanding how human corpora cavernosal tissue responds to SHH treatment is critical for clinical translation of SHH PA to ED patients. CONCLUSION: Corpora cavernosal smooth muscle from all ED patients responded to SHH treatment with increased growth. Stupp, SI. Sonic Hedgehog Signaling in Primary Culture of Human Corpora Cavernosal Tissue From Prostatectomy, Diabetic, and Peyronie's Patients. J Sex Med 2022;19:1228-1242.

Caspase Signaling in ED Patients and Animal Models.

BACKGROUND: Current treatments for erectile dysfunction (ED) are ineffective in prostatectomy and diabetic patients due to cavernous nerve (CN) injury, which causes smooth muscle apoptosis, penile remodeling, and ED. Apoptosis can occur via the intrinsic (caspase 9) or extrinsic (caspase 8) pathway. AIM: We examined the mechanism of how apoptosis occurs in ED patients and CN injury rat models to determine points of intervention for therapy development. METHODS AND OUTCOMES: Immunohistochemical and western analyses for caspase 3-cleaved, caspase-8 and caspase-9 (pro and active forms) were performed in corpora cavernosal tissue from Peyronie's, prostatectomy and diabetic ED patients (n = 33), penis from adult Sprague Dawley rats that underwent CN crush (n = 24), BB/WOR diabetic and control rats (n = 8), and aged rats (n = 9). RESULTS: Caspase 3-cleaved was observed in corpora cavernosa from Peyronie's patients and at higher abundance in prostatectomy and diabetic tissues. Apoptosis takes place primarily through the extrinsic (caspase 8) pathway in penis tissue of ED patients. In the CN crushed rat, caspase 3-cleaved was abundant from 1-9 days after injury, and apoptosis takes place primarily via the intrinsic (caspase 9) pathway. Caspase 9 was first observed and most abundant in a layer under the tunica, and after several days was observed in the lining of and between the sinuses of the corpora cavernosa. Caspase 8 was initially observed at low abundance in the rat corpora cavernosa and was not observed at later time points after CN injury. Aged and diabetic rat penis primarily exhibited intrinsic mechanisms, with diabetic rats also exhibiting mild extrinsic activation. CLINICAL TRANSLATION: Knowing how and when to intervene to prevent the apoptotic response most effectively is critical for the development of drugs to prevent ED, morphological remodeling of the corpora cavernosa, and thus, disease management. STRENGTHS AND LIMITATIONS: Animal models may diverge from the signaling mechanisms observed in ED patients. While the rat utilizes primarily caspase 9, there is a significant flux through caspase 8 early on, making it a reasonable model, as long as the timing of apoptosis is considered after CN injury. CONCLUSIONS: Apoptosis takes place primarily through the extrinsic caspase 8 dependent pathway in ED patients and via the intrinsic caspase 9 dependent pathway in commonly used CN crush ED models. This is an important consideration for study design and interpretation that must be taken into account for therapy development and testing of drugs, and our therapeutic targets should ideally inhibit both apoptotic mechanisms. Martin S, Harrington DA, Ohlander S, et al. Caspase Signaling in ED Patients and Animal Models. J Sex Med 2021;18:711-722.

Functional characterization of ATF1, GREM2 AND WNT10B variants associated with tooth agenesis.

OBJECTIVE: To determine the functional effects of ATF1, WNT10B and GREM2 gene variants identified in individuals with tooth agenesis (TA). SETTINGS AND SAMPLE POPULATION: Stem cells from human exfoliated deciduous teeth (SHED) were used as an in vitro model system to test the effect of TA-associated variants. MATERIALS AND METHODS: Plasmid constructs containing reference and mutant alleles for ATF1 rs11169552, WNT10B rs833843 and GREM2 rs1414655 variants were transfected into SHED for functional characterization of variants. Allele-specific changes in gene transcription activity, protein expression, cell migration and proliferation, and expression of additional tooth development genes (MSX1, PAX9 and AXIN2) were evaluated. Data analyses were performed using Student's t-test. P-values ≤ .05 were considered statistically significant. RESULTS: Mutant variants resulted in significantly decreased transcriptional activity of respective genes (P < 0.05), although no changes in protein localization were noted. Expression of MSX1 was significantly decreased in ATF1- and GREM2-mutant cells, whereas PAX9 or AXIN2 mRNA expression was not significantly altered. Mutant WNT10B had no significant effect on the expression of additional TA genes. ATF1- and GREM2-mutant cells presented increased cell migration. Cell proliferation was also affected with all three mutant alleles. CONCLUSIONS: Our results demonstrate that ATF1, WNT10B and GREM2 mutant alleles have modulatory effects on gene/protein function that may contribute to TA.

Peptide amphiphile nanofiber hydrogel delivery of Sonic hedgehog protein to the penis and cavernous nerve suppresses intrinsic and extrinsic apoptotic signaling mechanisms, which are an underlying cause of erectile dysfunction.

Erectile dysfunction (ED) is a common and debilitating condition with high impact on quality of life. An underlying cause of ED is apoptosis of penile smooth muscle, which occurs with cavernous nerve injury, in prostatectomy, diabetic and aging patients. We are developing peptide amphiphile (PA) nanofiber hydrogels as an in vivo delivery vehicle for Sonic hedgehog protein to the penis and cavernous nerve to prevent the apoptotic response. We examine two important aspects required for clinical application of the biomaterials: if SHH PA suppresses intrinsic (caspase 9) and extrinsic (caspase 8) apoptotic mechanisms, and if suppressing one apoptotic mechanism forces apoptosis to occur via a different mechanism. We show that SHH PA suppresses both caspase 9 and 8 apoptotic mechanisms, and suppressing caspase 9 did not shift signaling to caspase 8. SHH PA has significant clinical potential as a preventative ED therapy, by management of intrinsic and extrinsic apoptotic mechanisms.

Effect of surface preparation and light curing on penetration of silver particles from 38% silver diamine fluoride in dentin of primary teeth: An in vitro evaluation.

PURPOSE: To evaluate the effect of light cure, as well as various dentin surface treatment approaches, on the penetration depth of silver precipitating from 38% silver diamine fluoride into primary dentin tubules. METHODS: The occlusal dentin surfaces of 42 non-carious primary molars were exposed and then sectioned into halves bucco-lingually. The halves from each tooth pair were randomly split in two mega-groups, and each mega-group was divided randomly as follows into six experimental groups: prepared by either carbide bur (G1, G2), ceramic bur (G3, G4), or erbium laser (G5, G6). SDF was then applied to all prepared surfaces, and finally even-numbered groups (G2, G4, G6) were light cured. One mega-group was assigned to quantitative evaluation of silver penetration depth along the axial wall, and the other mega-group was reserved for qualitative observation of relative silver distribution on the occlusal surface, both via scanning electron microscope. RESULTS: No significant difference was observed in silver penetration depth between light cure and non-light cure groups (P= 0.8908). There was a statistically significant association between tooth preparation method and depth of silver penetration (P< 0.000001); laser-treated groups had significantly deeper silver penetration (1,148.9 µm G5, 1160.4 µm G6) than carbide bur (P< 0.05; 184.7 µm G1, 301.8 µm G2) or ceramic bur (P< 0.05; 184.1 µm G3, 131.0 µm G4) groups. A significant difference (P< 0.05) was noted in percentage occlusal surface coverage of particles between laser (51.4% G5, 35.8% G6) and carbide groups (21.1% G1, 19.3% G2). Light cure had no significant effect on the depth of silver penetration from 38% SDF in the dentin of primary teeth. Laser preparation resulted in deeper silver penetration than carbide or ceramic bur. CLINICAL SIGNIFICANCE: Exposure of 38% silver diamine fluoride-treated dentin to light cure did not affect the depth of penetration of silver particles into the dentin tubules of primary teeth. Rather, tooth preparation approaches that reduce the smear layer, like laser ablation, resulted in the deepest penetration of silver into the tubules. Clinical application of these findings will depend on scenario and treatment aim.

The Environmental Health Impact of Hurricane Katrina on New Orleans.

Hurricane Katrina caused unprecedented flood damage to New Orleans, Louisiana, and has been the costliest hurricane in US history. We analyzed the environmental and public health outcomes of Hurricane Katrina by using Internet searches to identify epidemiological, sociodemographic, and toxicological measurements provided by regulatory agencies.Atmospheric scientists have now warned that global warming will increase the proportion of stronger hurricanes (categories 4-5) by 25% to 30% compared with weaker hurricanes (categories 1-2).With the new $14.6 billion Hurricane Storm Damage Risk Reduction System providing a 100-year storm surge-defensive wall across the Southeast Louisiana coast, New Orleans will be ready for stronger storms in the future.

Optimization of Sonic Hedgehog Delivery to the Penis from Self-Assembling Nanofiber Hydrogels to Preserve Penile Morphology after Cavernous Nerve Injury.

Erectile dysfunction (ED) is a significant medical condition, with high impact on patient quality of life. Current treatments are minimally effective in prostatectomy, diabetic and aging patients due to injury to the cavernous nerve (CN); loss of innervation causes extensive smooth muscle (SM) apoptosis, increased collagen and ED. Sonic hedgehog (SHH) is a critical regulator of penile SM. We developed a self-assembling peptide amphiphile (PA) nanofiber hydrogel for extended release of SHH protein to the penis after CN injury, to suppress SM apoptosis. In this study we optimize the animal model, SHH concentration, duration of suppression, and location of delivery, to maximize SM preservation. SHH treatment suppressed apoptosis and preserved SM 48%. Increased SHH duration preserved SM 100%. Simultaneous penis/CN delivery increased SM 127%. Optimization of SHH PA delivery is essential for clinical translation to ED patients, and the PA vehicle has wide applicability as an in vivo delivery tool.

Sonic hedgehog regulation of human rhabdosphincter muscle:Potential implications for treatment of stress urinary incontinence.

AIMS: Rhabdosphincter (RS) muscle injury occurs during prostatectomy, and is a leading cause of stress urinary incontinence (SUI). Current SUI treatments engender significant side effects, which negatively impact patient quality of life. Thus an unmet need exists to develop novel RS regeneration methods. We have shown that Sonic hedgehog (SHH) is a critical regulator of penile smooth muscle, and we have developed novel peptide amphiphile nanofiber hydrogel delivery of SHH protein to the penis to regenerate smooth muscle after prostatectomy induced injury. If similar SHH signaling mechanisms regulate RS muscle homeostasis, this innovative technology may be adapted for RS regeneration post-prostatectomy. We examine the SHH pathway in human RS muscle. METHODS: Human RS obtained during radical cystoprostatectomy (n = 13), underwent SHH pathway analysis. Primary cultures were established (n = 5), and RS cells were treated with SHH protein, SHH inhibitor, or PBS (control). Immunohistochemical analysis for SHH pathway, skeletal muscle actin, and trichrome stain were performed. RS growth was quantified at 3 and 6 days. RESULTS: SHH, it is receptors patched and smoothened, and transcriptional activators, GLI proteins, were identified in human RS muscle. At 3 and 6 days, RS cells increased 62% and 78% (P = 0.0001) with SHH treatment and decreased 40% (P = 0.0001) and 18% (P = 0.039) with SHH inhibition. CONCLUSIONS: The SHH pathway was identified in human RS. RS growth increased with SHH treatment, indicating intervention may be possible to enhance RS regeneration, and impact SUI. Peptide amphiphile delivery of SHH may be applicable for RS regeneration and SUI prevention.

Peptide amphiphile delivery of sonic hedgehog protein promotes neurite formation in penile projecting neurons.

Erectile dysfunction (ED) critically impacts quality of life in prostatectomy, diabetic and aging patients. The underlying mechanism involves cavernous nerve (CN) damage, resulting in ED in 80% of prostatectomy patients. Peptide amphiphile (PA) nanofiber hydrogel delivery of sonic hedgehog (SHH) protein to the injured CN, improves erectile function by 60% at 6 weeks after injury, by an unknown mechanism. We hypothesize that SHH is a regulator of neurite formation. SHH treatment promoted extensive neurite formation in uninjured and crushed CNs, and SHH inhibition decreased neurites >80%. Most abundant neurites were observed with continuous SHH PA treatment of crushed CNs. Once induced with SHH, neurites continued to grow. SHH rescued neurite formation when not given immediately. SHH is a critical regulator of neurite formation in peripheral neurons under uninjured and regenerative conditions, and SHH PA treatment at the time of injury/prostatectomy provides an exploitable avenue for intervention to prevent ED.

Exploring parent-reported barriers to supporting their child's health behaviors: a cross-sectional study.

BACKGROUND: Parents can influence the health behaviors of their children by engaging in supportive behaviors (e.g., playing outside with their child, limiting recreational screen time). How, and the extent to which parents engage in supportive behaviors may be influenced by perceived barriers. The purpose of this study is to explore whether the frequency, and types, of barriers to providing parental support are dependent on the type of child health behavior being supported (i.e., physical activity, recreational screen time reduction, healthy eating, and sleep). METHODS: Study participants were 1140 Ontario parents with at least one child under the age of 18 who completed a Computer Assisted Telephone Interview (CATI) survey about parental support behaviors. Open-ended responses about perceived barriers to parental support were coded, and aggregated to meta-categories adopted from the social-ecological model (i.e., individual child, individual parent, interpersonal, environmental). Freidman rank sum tests were used to assess differences across child behaviors. Wilcoxon rank sum tests with Bonferroni adjustments were used as a post hoc test for significant Freidman results. RESULTS: There were more barriers reported for supporting physical activity than for any other child behavior (ps < .01, As ≥ .53). Parents reported more parent level and environmental level barriers to supporting child physical activity versus other behaviors (ps < .001, As ≥ .55), child level barriers were more frequently reported for supporting healthy eating and sleep (ps < .001, As ≥ .57), and interpersonal barriers were more frequently reported for supporting recreational screen time reduction (ps < .001, As ≥ .52). Overall, parents reported more child and parent level barriers versus interpersonal and environmental barriers to supporting child health. CONCLUSIONS: Parents experience a variety of barriers to supporting their children's health behaviors. Differences in types of barriers across child health behaviors emerged; however, some frequently reported barriers (e.g., child preferences) were common across behaviors. Interventions promoting parental support should consider strategies that can accommodate parents' busy schedules, and relate to activities that children find enjoyable. Creating supportive environments that help facilitate support behaviors, while minimizing parent level barriers, may be of particular benefit. Future research should explore the impact of barriers on parental support behaviors, and effective strategies for overcoming common barriers.

Do parents' support behaviours predict whether or not their children get sufficient sleep? A cross-sectional study.

BACKGROUND: Sleep is an essential component of healthy cognitive and physical development. Lack of sleep may put children at risk for a variety of mental and physical health outcomes, including overweight, obesity and related chronic diseases. Given that children's sleep duration has decreased in recent decades, there is a need to understand the determinants of child sleep, including the role of parental support behaviours. This study aims to determine the relative contribution of different types of parental support behaviours for predicting the likelihood that children meet recently established Canadian sleep guidelines. METHODS: Data were collected using Computer Assisted Telephone Interviews (CATI) of parents or guardians with at least one child under the age of 18 living in Ontario, Canada. To align with sleep guidelines, parents included in this analysis had at least one child between 5 and 17 years of age (n = 1622). Two multivariable logistic regression models were built to predict whether or not parents reported their child was meeting sleep guidelines - one for weekday sleep and another for sleep on weekends. Independent variables included parent and child age and gender, motivational and regulatory parental support behaviours, and socio-demographic characteristics. RESULTS: On weekdays, enforcing rules about child bedtime was a significant positive predictor of children meeting sleep guidelines (OR: 1.59; 95% CI: 1.03-2.44); while encouraging the child to go to bed at a specific time was a significant negative predictor of child meeting sleep guidelines (OR: 0.29; 95% CI: 0.13-0.65). On weekends, none of the parental support behaviours contributed significantly to the predictions of child sleep. For both weekdays and weekends, the child's age group was an important predictor of children meeting sleep guidelines. CONCLUSIONS: The contribution of parental support behaviours to predictions of children meeting sleep guidelines varied with the type of support provided, and weekend versus weekday sleep. While only enforcing bedtime rules on weekdays contributed to children meeting sleep guidelines, the importance of children getting a good night's sleep, and the capacity of parents to help them do so, should be emphasized in public health efforts promoting healthy child development.