Intracavernosal pressure characteristics analyzed by power spectral density for identifying the phase of penile erection.

Knowledge of the time course of penile erection is very important to understanding erection physiology. The changes in the intracavernosal pressure (ICP) and the different phases of an erection are pivotal to the ability to produce and maintain a rigid penile erection. This study investigated an objective and low-invasiveness method for identifying different erection phases based on an innovative ICP analysis technique. Blood infuses into the corpora cavernosa and causes the ICP to increase. The ICP usually exhibits tiny oscillations at the frequency of the heartbeat when it increases from diastole to systole. The characteristic oscillation amplitudes corresponding to the period when the full and rigid erection phases begin can be extracted by power spectral density analysis. The reliability and accuracy of the proposed method was verified by the Bland-Altman graphs indicating a good agreement with the existing method that compares the ICP with the arterial pressure. Moreover, all of the intraclass correlation coefficient values were close to 1.00, with the lower limit of the 95% confidence interval exceeding 0.75. The described novel objective and low-invasiveness method can therefore be used for identifying the full and rigid erection phases of the penis in urological investigations during different erection phases.

Direct repair vs. overlapping sphincter repair: a randomized, controlled trial.

PURPOSE: The aim of this study was to compare the results of two surgical techniques (direct end-to-end vs. overlapping) of delayed repair of a localized anterior defect of external anal sphincter after an obstetric trauma. METHODS: During a five-year period, 23 patients were randomly assigned to direct end-to-end repair (n = 12) or overlapping sphincter repair (n = 11), using 2-0 PDS sutures. Two patients from each group had an internal anal sphincter defect that also was repaired. All patients had a normal pudendal nerve terminal motor latency preoperatively. Evaluations included endoanal ultrasound, anorectal manometry, and neurophysiologic evaluation. Continence was assessed by the Cleveland Clinic Continence Score (0-20; 0, perfect continence; 20, complete incontinence). RESULTS: The two groups were comparable with regard to age (median, 45 years), past history of sphincter repair (n = 2), and posterior vaginal repair. There was no major morbidity. The wound-healing rate was identical between the two groups. However, of the patients undergoing overlapping repair, two had fecal impaction, and one had a urinary retention. Median preoperative continence score was 17 in both the direct-repair group (score, 8-20) and the overlap group (score, 7-20). At a median follow-up of 18 months, the improvement in continence was similar between the two surgical groups, with a median continence score of 3, respectively. In both surgical groups there was a significant and similar improvement in maximum squeeze pressure and in the functional anal canal length postoperatively (P < 0.05), but the mean resting pressure was relatively unchanged. In the overlap group, one patient developed a unilaterally prolonged pudendal nerve terminal motor latency that was persistent 22 months after surgery, and two patients had impaired fecal evacuation postoperatively. CONCLUSIONS: This randomized, controlled study suggests that the outcome is similar whether direct end-to-end or overlapping repair of a sphincter defect is performed. Overlapping repair might be associated with more difficulties with fecal evacuation and a prolonged pudendal nerve terminal motor latency postoperatively.

Faecal incontinence after lateral internal sphincterotomy is often associated with coexisting occult sphincter defects: a study using endoanal ultrasonography.

BACKGROUND: Troublesome faecal incontinence following a lateral internal sphincterotomy (LIS) is often attributed to faulty surgical techniques: division of excessive amount of internal sphincter or inadvertent injury to the external sphincter. The aim of the present paper was to assess the anatomic and physiological factors that may contribute to faecal incontinence following a technically satisfactory lateral internal sphincterotomy by a group of colorectal specialists. METHODS: Fourteen patients (nine women, five men; median age: 38 years; range: 23-52 years) who developed troublesome postoperative faecal incontinence were evaluated by clinical assessment, endoanal ultrasonography and anorectal physiological studies (manometry, pudendal nerve terminal motor latency) by two independent observers. The Cleveland Clinic continence score (0-20; 0, perfect continence; 20, complete incontinence) was used to quantify the severity of faecal incontinence. Fourteen continent subjects after a LIS (nine female patients, five male patients; median age: 36 years; range: 20-44 years) were also evaluated as 'continent' controls (continence score

Prolonged allograft survival in anti-CD4 antibody transgenic mice: lack of residual helper T cells compared with other CD4-deficient mice.

BACKGROUND: Investigations of the role of CD4 T lymphocytes in allograft rejection and tolerance have relied on the use of mouse models with a deficiency in CD4 cells. However, in mice treated with depleting monoclonal antibody (mAb) and in MHC class II knockout (KO) mice, there are residual populations of CD4 cells. CD4 KO mice had increased CD4- CD8-TCRalphabeta+ helper T cells, and both strains of KO mice could reject skin allografts at the normal rate. In this study, transgenic mice with no peripheral CD4 cells were the recipients of skin and heart allografts. Results were compared with allograft survival in CD4 and MHC class II KO mice. METHODS: GK5 (C57BL/6 bml mice transgenic for a chimeric anti-CD4 antibody) had no peripheral CD4 cells. These mice, and CD4 and class II KO mice, received BALB/c or CBA skin or cardiac allografts. Some GK5 mice were treated with anti-CD8 mAb to investigate the role of CD8 cells in rejection. CD4 and CD8 cells were assessed by FACS and immunohistochemistry. RESULTS: BALB/c skin on GK5 mice had a mean survival time +/- SD of 24+/-6 days, compared with 9+/-2 days in wild-type mice. Anti-CD8 mAb prolonged this to 66+/-7 days. BALB/c skin survived 10+/-2 days on class II KO and 14+/-2 days on CD4 KO, both significantly less than the survival seen on GK5 recipients (P<0.001). BALB/c hearts survived >100 days in GK5 recipients and in wild-type recipients treated with anti-CD4 mAb at the time of grafting, in contrast to a mean survival time of 10+/-2 days in untreated wild-type mice. Immunohistochemistry revealed that long-term surviving heart allografts from the GK5 recipients had CD8 but no CD4 cellular infiltrate. These hearts showed evidence of transplant vasculopathy. CONCLUSIONS: The GK5 mice, with a complete absence of peripheral CD4 cells, provide the cleanest available model for investigating the role of CD4 lymphocytes in allograft rejection. Prolonged skin allograft survival in these mice compared with CD4 and MHC class II KO recipients was clearly the result of improved CD4 depletion. Nevertheless, skin allograft rejection, heart allograft infiltration, and vascular disease, mediated by CD8 cells, developed in the absence of peripheral CD4 T cells.

Anorectal physiologic testing for bowel dysfunction in patients with spinal cord lesions.

PURPOSE: Bowel dysfunction is common in patients with spinal cord lesions. This study aims to determine whether there are any discriminatory findings at anorectal physiologic testing in patients with spinal cord lesions. METHODS: Twelve consecutive patients (6 females) with significant spinal cord lesions who had mixed symptoms of constipation, fecal impaction, and fecal incontinence were evaluated by perfusion manometry and pudendal nerve terminal motor latency. None of the patients had a sphincter defect as evaluated by endoanal ultrasonography. RESULTS: The median age was 54 (range, 40-87) years. Eight (67 percent) of them had had traumatic spinal cord injuries. Other spinal cord lesions included spina bifida, syringomyelia, arachnoid cyst, and spinal cord ischemia after abdominal aortic aneurysm repair. In patients with spinal cord lesions, the mean (range) resting anal canal pressure and maximum squeeze anal canal pressure were 46 (10-100) mmHg and 76 (30-120) mmHg respectively compared with 62 (50-70) mmHg, and 138 (100-180) mmHg, respectively, in healthy controls. Eleven (92 percent) patients had prolonged pudendal nerve terminal motor latency (9 bilateral and 2 unilateral) whereas rectoanal inhibitory reflex was abolished in all 9 patients tested. CONCLUSIONS: Spinal patients with severe bowel symptoms tended to have lower anal canal pressures than healthy controls. Pudendal netropathy and impaired rectoanal inhibitory reflex are common and may be important in the pathogenesis of bowel dysfunction in patients with spinal cord lesions.

Modified technique for kidney transplantation in mice.

This study describes a new method for joining the donor ureter to the recipient bladder during mouse kidney transplantation. The donor left kidney was harvested using methods previously published, except that bladder tissue was not harvested with the end of the ureter. The recipient left kidney was removed and the donor kidney was attached using end-to-side anastomosis. The recipient bladder was pierced with a 21-gauge needle allowing curved forceps to be inserted through the bladder, to pull through the ureter, and the periuretal tissue was stitched to the exterior wall of the bladder. The donor ureter was allowed to retract inside the bladder. Following a right nephrectomy, grafts were monitored by blood serum creatinine and urea. With a technical success rate of 83%, this technique reduced donor harvest time by 20 minutes and ureter attachment time by 15 minutes making it the best method available for mouse kidney transplantation.

Assessment of peripheral tolerance in anti-CD4 treated C57BL/6 mouse heart transplants recipients.

The study was designed to compare second heart and skin grafts and in vitro assays as a means of assessing peripheral tolerance in C57BL/6 mice. Vascularized heterotopic BALB/c hearts were placed in C57BL/6 recipients treated with anti-CD4, GK1.5 (1 mg total per 20 g mouse i.p. on days 0, 1, 2, 3). Those mice in which hearts survived for >60 days were challenged with donor and third-party (CBA) skin grafts or with second heart grafts, of donor or third-party origin, attached to the carotid artery and jugular vein. In vitro alloreactivity was assessed by mixed lymphocyte reactions (MLR) and cell mediated lympholysis (CML) using recipient spleen cells. Parenchymal damage, cellular infiltration and vascular disease were assessed from the histology of long-term allografts and isografts. Allografts in untreated recipients were rapidly rejected while isografts survived > 100 days. Primary allografts in anti-CD4 treated recipients also survived > 100 days, as did donor strain secondary heart transplants given at >60 days after the first graft. Third-party hearts were rapidly rejected, as were donor and third-party skin grafts placed on recipients with long-term allografts. These recipients showed low MLR response to both donor and third-party stimulators and donor-specific suppression of CML at 60 days post graft. Long-surviving heart allografts all showed evidence of parenchymal damage and vascular intimal thickening. Thus in the BALB/c to C57BL/6 donor-recipient strain combination, hearts, but not skin grafts, could be used to demonstrate peripheral tolerance, which seemed to be both organ and major histocompatibility complex (MHC) specific. Despite long survival, BALB/c hearts all showed evidence of parenchymal damage and vascular intimal thickening, a sign of chronic rejection.

Idarubicin-145-2C11-F(ab')2 promotes peripheral tolerance and reduces chronic vascular disease in mouse cardiac allografts.

In order to reduce the toxic effects of the T cell activating anti-CD3 monoclonal antibody, 145-2C11, F(ab')2 fragments were prepared by pepsin digestion. These fragments were then used as non-immunosuppressive carriers for the cytotoxic drug idarubicin (IDA), to reduce toxicity of both the monodonal antibodies (mAb) and the drug and to increase the specificity of drug delivery. The IDA-145-2C11 F(ab')2 immunoconjugate was tested for specificity by fluorometry. 145-2C11 intact antibody, 145-2C11 F(ab')2 and IDA conjugates of the antibody and F(ab')2 were used to treat CBA recipients of BALB/c vascularized cardiac allografts. Mice with hearts surviving >100 days were challenged with donor and third party (C57BL/6) skin grafts. Although both antibody and F(ab')2 blocked the binding of 145-2C11-FITC to CBA spleen cells, only the intact antibody caused sustained depletion of CD3 cells in vivo. 145-2C11 F(ab')2 blocked cell surface CD3 within 30 min, but was cleared in 24 h without depletion of CD3 cells from the spleen. In BALB/c to CBA cardiac allografts (rejected in 12-17 days), IDA-145-2C11 F(ab')2 (0.2 mg/20 g mouse i.p. at the time of transplantation) induced >100 days' allograft survival and specific tolerance, in contrast to the equivalent dose of 145-2C11 F(ab')2 (mean survival 25 days). Hearts from IDA-145-2C11 F(ab')2-treated mice at >100 days showed decreased cellular infiltration and less chronic vascular disease than long-surviving hearts from mice treated with an alternative antibody, KT3. Thus, F(ab')2 prepared from 145-2C11 provided a suitable CD3-specific, nonimmunosuppressive carrier for IDA. This immunoconjugate was more effective against both acute and chronic rejection than other conjugates or whole antibody. IDA-145-2C11 F(ab')2 is an effective, nontoxic tolerogen in the mouse cardiac allograft model.

Effect of IL-4 deletion on cardiac allograft survival in the BALB/c to 129Sv x C57BL/6 strain combination.

To investigate the effect of IL-4 deletion on cardiac allograft survival, vascularized BALB/c cardiac allografts were placed in C57BL/6, 129Sv x C57BL/6 (IL-4 +/+) or 129Sv x C57BL/6 IL-4 knockout mice (IL-4-/-). Untreated recipients rejected allografts in < 15 days while isografts survived indefinitely (> 100 days). Treatment with anti-CD4 (GK1.5) for 4 days at the time of allografting increased mean survival to > 100 days in C57BL/6, 90+/-16 days in 129Sv x C57BL/6 (IL-4 +/+) and 68 +/- 36 days in 129Sv x C57BL/6 (IL-4-/-) recipients. Although there was a trend towards shorter survival times in the IL-4-/- mAb-treated mice, survival in the three recipient groups was not significantly different (P = 0.07). A 30-day course of anti-CD4 did not further prolong BALB/c heart survival. All long-surviving hearts had histological evidence of parenchymal damage and transplant vascular disease. None of these recipients developed antigen-specific tolerance, since both donor and third party skin graft challenges were rejected when challenged at > 60 days post-graft and all primary grafts failed by 120 days. Thus the effects of IL-4 deletion were subtle and were seen only with low doses of immunosuppression in this high responder strain combination.

Idarubicin-anti-CD3: a new immunoconjugate that induces alloantigen-specific tolerance in mice.

BACKGROUND: In testing new anti-CD3 agents for transplantation tolerance induction, an anti-CD3 monoclonal antibody was used as a carrier for the cytotoxic drug idarubicin (IDA). METHODS: Anti-CD3 (KT3) was covalently coupled with IDA, producing the IDA-KT3 immunoconjugate, which was tested for specificity by fluorometry and for inhibition of proliferation of CD3+ E3 cells ([3H]thymidine uptake). KT3 and IDA-KT3 were used to treat CBA recipients of BALB/c vascularized cardiac allografts. Mice with hearts surviving >100 days were challenged with donor and third-party (C57BL/6) skin. RESULTS: Conjugation to IDA did not reduce binding of KT3 to E3 cells, although the toxicity of IDA was reduced by conjugation. In BALB/c to CBA cardiac allografts (rejected in 12-17 days), both KT3 and IDA-KT3 (0.25-0.5 mg/20 g mouse i.p. at the time of transplantation) induced tolerance. Hearts survived >100 days and skin graft challenge showed indefinite survival of donor grafts but not third-party grafts. KT3 was less toxic, as measured by tumor necrosis factor-a release and blood glucose levels, than equivalent dosages of 145-2C11. At lower dosages (0.1 mg/20 g mouse), KT3-treated animals rejected BALB/c allografts in 15 to 19 days, but IDA-KT3 induced long survival (>100 days) and donor-specific tolerance in 5 of 6 mice. CONCLUSIONS: Coupling IDA to anti-CD3 reduced the in vivo toxicity of IDA and improved the immunosuppressive performance of KT3, reducing the side effects seen with other anti-CD3 agents. IDA-KT3 is a new, effective, nontoxic tolerogen in this donor-recipient combination.

Interleukin (IL) 4, the cytokine that isn't there: reactivity of IL-4 antibodies with cells in IL-4 -/- mice.

We analyzed cytokine expression in recipient spleens and cardiac allografts placed in mice that were unable to synthesize interleukin (IL)-4 due to disruption of the IL-4 gene (IL-4 -/-) and in wild-type (IL-4 +/+) mice. Polyclonal BL-4P and monoclonal 11B11, 1D11, and 24G2 anti-IL-4 antibodies were used to detect cell-surface and cytoplasmic antigens in sections of frozen tissue. All of the antibodies were found to react with non-IL-4 determinants associated with graft-infiltrating cells, and BL-4P, 1D11, and 24G2 bound to cells and connective tissue in the spleens of IL-4 -/- mice. The IL-4-producing cell line, X63Ag8-653 (X63), was used as a positive control for IL-4 staining and to test the ability of recombinant IL-4 to block the binding of antibodies to IL-4.

Increased expression of IL-4 and IL-10 and decreased expression of IL-2 and interferon-gamma in long-surviving mouse heart allografts after brief CD4-monoclonal antibody therapy.

In a mouse model for vascularized heart transplantation, CBA recipients of BALB/c hearts were treated with 0.25 mg of anti-CD4 (GK1.5) given intraperitoneally on the day of grafting and on days 1, 2, and 3 thereafter. This reduced splenic CD4+ cells to < 1% and all grafts survived > 100 days, compared with 8-10 days in untreated recipients. Despite recovery of the CD4+ cells after day 21, mice did not reject donor-type skin grafts at > 30 days, but rapidly rejected third-party skin, showing alloantigen-specific tolerance. The surviving heart grafts had significant mononuclear cell infiltration at time points from 7 to 100 days after transplantation. In the normal rejection process, where extensive myocyte necrosis was seen at 7 days, graft-infiltrating T cells produced IL-2 and IFN-gamma. These cells responded in vitro to IL-2 and displayed donor-specific CTL activity. In contrast, cells from CD4-mAb-treated hearts did not show significant growth in IL-2 or kill donor cells in CTL assays. In these nonrejecting hearts, immunohistology showed a diffuse infiltrate of T cells and macrophages by day 3. The allograft infiltrate increased rapidly thereafter in both rejecting and nonrejecting grafts, peaking at day 6-7 in rejecting grafts, when CD4+, CD8+, and IL-2R+ cells were present, with expression of IL-2, IFN-gamma, and IL-4, but only trace levels of IL-10. From 14 to 100 days, nonrejecting allografts showed a characteristic cytokine profile of dense IL-4 and IL-10 expression on intragraft leukocytes and endothelial cells, with low levels of IL-2 and IFN-gamma. This cytokine profile, characteristic of Th2 responses, was seen in all nonrejecting grafts and was not present in rejecting grafts. Allograft tolerance can studied by examination of the functions and cytokine profile of the cells within the graft, and tolerance develops in the presence of a Th2 response within the graft.

Photodynamic therapy in the treatment of subcutaneously implanted human bladder tumour.

Photodynamic therapy (PDT) is an experimental treatment modality for malignant tumours. The effect of PDT with haematoporphyrin derivative (HpD) was studied using a human bladder tumour (BL-17) which was implanted subcutaneously (s.c.) into immunodeficient Balb/c nude mice. This model is only suitable for short-term investigation of PDT because of the high mortality that arises due to the immune deficiency of the animals. In a short-term observation (2 weeks post-treatment), HpD sensitized PDT was effective in the control of tumour growth, with 71% of tumours cured. The effect of PDT was found to be highly dependent on doses of HpD and/or the activating laser light. The comparison of PDT effects of the gold metal vapour laser (GMVL) and argon ion pumped dye laser (AIPDL) indicated that no significant difference exists between these two different laser sources for PDT. The irradiation with laser light alone and the administration of HpD alone had no significant effect on tumour growth.