Inhibition of DNA methylation during chronic obstructive bladder disease (COBD) improves function, pathology and expression.

Partial bladder outlet obstruction due to prostate hyperplasia or posterior urethral valves, is a widespread cause of urinary dysfunction, patient discomfort and also responsible for immense health care costs. Even after removal or relief of obstruction, the functional and pathologic aspects of obstruction remain as a chronic obstructive bladder disease (COBD). Epigenetic changes, such as DNA methylation, contribute to the persistent character of many chronic diseases, and may be altered in COBD. We tested whether candidate genes and pathways and the pathophysiology of COBD were affected by a hypomethylating agent, decitabine (DAC). COBD was created in female Sprague-Dawley rats by surgical ligation of the urethra for 6 weeks, followed by removal of the suture. Sham ligations were performed by passing the suture behind the urethra. After removal of the obstruction or sham removal, animals were randomized to DAC treatment (1 mg/kg/3-times/week intraperitoneally) or vehicle (normal saline). Bladder function was non-invasively tested using metabolic cages, both one day prior to de-obstruction at 6 weeks and prior to sacrifice at 10 weeks. Residual volume and bladder mass were measured for each bladder. Bladders were examined by immunostaining as well as qPCR. The effects of DNA methyltransferase (DNMT)-3A knockout or overexpression on smooth muscle cell (SMC) function and phenotype were also examined in bladder SMC and ex vivo culture. Residual volumes of the DAC treated group were not significantly different from the NS group. Compared to COBD NS, COBD DAC treatment helped preserve micturition volume with a significant recovery of the voiding efficiency (ratio of the maximum voided volume/maximum bladder capacity) by one third (Fig. 1, p > 0.05). Brain-derived neurotrophic factor (BDNF) variants 1 and 5 were upregulated by COBD and significantly reduced by DAC treatment. Deposition of collagen in the COBD bladder was reduced by DAC, but gross hypertrophy remained. In bladder SMC, DNMT3A overexpression led to a loss of contractile function and phenotype. In bladders, persistently altered by COBD, inhibition of DNA-methylation enhances functional recovery, unlike treatment during partial obstruction, which exacerbates obstructive pathology. The underlying mechanisms may relate to the gene expression changes in BDNF and their effects on signaling in the bladder.

Urethral compliance in hypospadias operated by tubularized incised urethral plate (TIP) with and without a dorsal inlay graft: an experimental controlled study.

OBJECTIVE: Abnormal uroflowmetries are common after tubularized incised plate urethroplasties (TIP), perhaps due to low compliance. We hypothesized that (1) abnormal uroflowmetries after TIP might be caused by segmental lower compliance; (2) by adding a graft to the raw area in the incised plate (TIPG), compliance might be improved by preventing secondary intention healing of the dorsal incision. METHODS: A standardized penectomy was performed in 27 adult male rabbits: 9 normal non-operated controls (G1), 6 weeks after TIP (G2: n = 9) or TIPG (G3: n = 9). A standardized isolated segment (including the whole urethroplasty in G1 and G2) was progressively distended with air (1, 2 and 3 ml) in the 3 groups. The respective intraluminal pressures were measured with a tensiometer. RESULTS: Pressure measurements were feasible and reproducible for this model. Mean pressures tended to be higher in the experimental groups (G1: 59.7 mmHg vs. G2: 79.6 mmHg vs. G3: 100.1 mmHg for 1 ml injections; G1: 233.1 mmHg vs. G2: 241 mmHg vs. G3: 308.4 mmHg for 2 ml injections and G1: 457.3 mmHg vs. G2: 429 mmHg vs. G3: 520 mmHg for 3 ml injections) without reaching the statistical significance. CONCLUSION: In this model, the elasticity of the TIP or TIPG neourethras tended to be reduced when compared to controls. The placement of an inlay graft on the dorsal incised area did not increase the compliance. This model allows the measurement of segmental intraluminal urethral pressures generated by controlled air distension and may be a useful tool to evaluate the experimental urethroplasty models.

Extracellular matrix gene responses in a novel ex vivo model of bladder stretch injury.

PURPOSE: Congenital bladder outlet obstruction from either mechanical or functional causes often results in clinical bladder fibrosis. We tested the hypothesis that early molecular changes relevant to fibrosis occur in response to stretch injury of the bladder wall and that specific extracellular matrix receptors mediate some of these responses. Furthermore, we introduce a novel ex vivo model of bladder injury which has advantages over previously described in vivo bladder outlet obstruction models by uniquely interrogating molecular responses to bladder distention. MATERIALS AND METHODS: The bladders of Sprague Dawley rats were hydrodistended transurethrally, the ureters and bladder neck were ligated, and the whole bladder was excised and incubated in culture medium in the distended state. At fixed time-points control and stretch bladders were snap frozen, RNA was extracted, and semiquantitative reverse transcription polymerase chain reaction for collagens I, III and XII, and RHAMM (receptor for hyaluronic acid) messenger (m) RNA was performed to establish trends in stretch related gene expression. Bladder specimens were also subjected to routine histological evaluation. RESULTS: An average 3-fold reduction in collagen I mRNA expression was seen with 8 hours of static stretch (p <0.05). Bladder stretch increased collagen III mRNA levels approximately 2.5-fold (p <0.05). Whole bladder collagen XII and RHAMM mRNA were elevated as much as 5-fold (p <0.05) with stretch. Blocking RHAMM function significantly attenuated these matrix gene responses (p = 0.01 to 0.005). CONCLUSIONS: The ex vivo model of whole bladder stretch is viable and easily reproducible for the study of molecular pathophysiological mechanisms contributing to maladaptive bladder disease. Furthermore, collagen gene transcription is revealed to be rapidly responsive to stretch injury of the bladder. Intact RHAMM receptor function is involved in these responses. Elucidation of the intermediate steps in this response to injury may allow for the development of novel therapeutic strategies which may prevent pathological matrix remodeling seen in clinical bladder disease.

Infant intersubjectivity: research, theory, and clinical applications.

We review research evidence on the emergence and development of active "self-and-other" awareness in infancy, and examine the importance of its motives and emotions to mental health practice with children. This relates to how communication begins and develops in infancy, how it influences the individual subject's movement, perception, and learning, and how the infant's biologically grounded self-regulation of internal state and self-conscious purposefulness is sustained through active engagement with sympathetic others. Mutual self-other-consciousness is found to play the lead role in developing a child's cooperative intelligence for cultural learning and language. A variety of preconceptions have animated rival research traditions investigating infant communication and cognition. We distinguish the concept of "intersubjectivity", and outline the history of its use in developmental research. The transforming body and brain of a human individual grows in active engagement with an environment of human factors--organic at first, then psychological or inter-mental. Adaptive, human-responsive processes are generated first by interneuronal activity within the developing brain as formation of the human embryo is regulated in a support-system of maternal tissues. Neural structures are further elaborated with the benefit of intra-uterine stimuli in the foetus, then supported in the rapidly growing forebrain and cerebellum of the young child by experience of the intuitive responses of parents and other human companions. We focus particularly on intrinsic patterns and processes in pre-natal and post-natal brain maturation that anticipate psychosocial support in infancy. The operation of an intrinsic motive formation (IMF) that developed in the core of the brain before birth is evident in the tightly integrated intermodal sensory-motor coordination of a newborn infant's orienting to stimuli and preferential learning of human signals, by the temporal coherence and intrinsic rhythms of infant behaviour, especially in communication, and neonates' extraordinary capacities for reactive and evocative imitation. The correct functioning of this integrated neural motivating system is found to be essential to the development of both the infant's purposeful consciousness and his or her ability to cooperate with other persons' actions and interests, and to learn from them. The relevance of infants' inherent intersubjectivity to major child mental health issues is highlighted by examining selected areas of clinical concern. We review recent findings on postnatal depression, prematurity, autism, ADHD, specific language impairments, and central auditory processing deficits, and comment on the efficacy of interventions that aim to support intrinsic motives for intersubjective communication when these are not developing normally.

Self/other organization in human psychological development.

Evidence from the evolution of human cultural behavior and learning, embryology and genetics of the brain, and the behavior of human infants indicates that the critical and uniquely human motives for cooperative imagination and joint interest in objects and tasks are determined by expression of genes and epigenetic neural systems elaboration long before birth, along with essential peripheral organs of perception and motor expression that will serve in communication by rhythmic facial, vocal, gestural, and body movement signals. These cerebral motives continue to exercise their influence on neural development and behavior throughout life, transforming the behaviors of the developing individual through a succession of phases to which other individuals and cultural institutions are constrained to adapt. We discuss the theory of innate intersubjectivity and relate it to the hypothesis of an Innate Motive Formation that emerges in brain development as regulator of morphogenesis in neural systems, and that continues to function, postnatally, as generator of motives and emotions by which human contacts and relationships are regulated. We suggest that differentiates of the primary motive formation in the embryo brain later serve to generate intelligent exploration of the objective environment and the emergence of an additional dialogic mechanism that represents the self-subject as a partner for an other-subject, intersubjectively. Intersubjective communication in infancy leads, through systematic age-related transformations of the brain and behavior, to preverbal mimetic negotiation of cooperative awareness and joint task performance. Finally we discuss, in relation to this theory, interpretations of faulty communication and development at different stages of the life cycle that result from maternal postnatal depression, autism, premature birth and schizophrenia.