The animals were housed in individual cages, under a 12-hour light-dark regimen and humidity and temperature control.
Surgery was performed on animals anesthetized with diazepam, ketamine, and medetomidine.
After midline incision following the vesical ligament, we found the bladder. The bladder will be exteriorized and by a cystocath we will misured its capacity. After a 0,5 x 0,5 cm specimen of bladder will be removed and replaced with o 0,5 x 0,5 cm “2D” scaffold of PCL (polycaproilatone)
Animals received 5% glucose solution after surgery, wetted food from the second day after surgery, and then were allowed free access to food. Postoperative analgesia and the antibiotic enrofloxacin were given and animals’ weight was recorded daily.
The animals were fed a normal standard diet with no restriction on food or water supply for 1 week before surgery.
31 Tissue engineering and tissue replacement with acellular grafts
The alternative approaches being pursued to find a practical and functional substitute for native bladder tissue can be grouped into the two broad categories of in vivo tissue regeneration and in vitro tissue engineering .
IN VIVO TISSUE REGENERATION USING ACELLULAR MATRICES
In this approach, which is intended to take advantage of the capability of the bladder to undergo rapid regeneration and repair after acute injury, a biomaterial implanted at the time of surgery becomes cellularized and eventually is assimilated into the tissues of the host bladder. Of all the synthetic and natural materials used to date, decellularized xenogeneic or allogeneic matrices prepared from the submucosa of small intestine and bladder have shown greatest promise.
We waiting for the experimental protocol is approved by the local Ethics Committee of the Ospedale Cardarelli . The animals will be treated and housed according to national and international regulations governing the use of animals in scientific research.
In 6 months 20 Wistar rats weighing between 210 and 270 g will be used.
The animals were housed in individual cages, under a 12-hour light-dark regimen and humidity and temperature control.
Surgery was performed on animals anesthetized with diazepam, ketamine, and medetomidine.
After midline incision following the vesical ligament, we found the bladder. The bladder will be exteriorized and by a cystocath we will misured its capacity. After a 0,5 x 0,5 cm specimen of bladder will be removed and replaced with o 0,5 x 0,5 cm “2D” scaffold of PCL (polycaproilatone)
Animals received 5% glucose solution after surgery, wetted food from the second day after surgery, and then were allowed free access to food. Postoperative
32 analgesia and the antibiotic enrofloxacin were given and animals’ weight was recorded daily.
The animals were fed a normal standard diet with no restriction on food or water supply for 1 week before surgery.
Before surgery, rats were fasted overnight and weighed.
On the first postoperative day, rats had free access to water and rodent diet.
The animals were killed by CO2 inhalation 7, and 15 days after surgery.
33
Conclusive remarks
The use of small-intestinal submucosa (SIS) and bladder acellular matrix (BAM) materials have been investigated in bladder augmentation studies in rodent, canine and porcine models. These studies suggest that both SIS and BAM are biocompatible, biodegradable and facilitate tissue-specific morphological and functional regeneration. Indeed, Kropp et al. reported that when SIS was used to augment the bladder after partial cystectomy in dogs, histologically normal bladder tissue regenerated within a month. A more pertinent question is whether the regenerated tissue acquires the functional properties of normal bladder tissue. On this point the evidence is contradictory, as the authors of some quantitative morphometric studies have reported marked graft contraction and decreased muscle content. There are several possible reasons to account for the progressive shrinkage of acellular grafts, including resorption of the matrix, smooth muscle contraction and graft fibrosis. Although the exact cause remains to be established, graft fibrosis is clearly undesirable as it would adversely affect the biomechanical properties and compliance of the regenerated tissue.
The obvious advantage of acellular matrices lies in their potential as an ‘off the shelf’ solution to reconstructive bladder surgery.
Unfortunately, the processes required to produce a marketable material, e.g.
chemical cross-linking and sterilization, may adversely affect the physical properties of the matrix.
The same processes may also deactivate matrix-bound growth factors and other signalling molecules responsible for the regenerative properties of the fresh material.
As yet, there are no published reports that any decellularized matrix has been used in a commercially available form for bladder reconstruction in clinical
34 urology. In the future, advances in the understanding of the physical and chemical factors responsible for urothelial and smooth muscle cell proliferation, migration, differentiation and function may be exploited in the development of so-called ‘smart’ biomaterials, in which
biologically active agents are incorporated into acellular natural or synthetic matrices.
35
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