In the human urothelium and suburothelium, intradetrusor botulinum neurotoxin type A does not induce apoptosis: preliminary results

Intradetrusor injections of botulinum neurotoxin type A (BoNTA) are emerging as the preferred second-line treatment for neurogenic and idiopathic overactive bladder (OAB). In animal experiments, intradetrusor BoNTA injections have been shown to cause apoptosis in the bladder urothelium and suburothelium but not the detrusor.

Complete continence after botulinum neurotoxin type A injections for refractory idiopathic detrusor overactivity incontinence: patient-reported outcome at 4 weeks

Objective improvement following intradetrusor injections of botulinum neurotoxin type A (BoNTA) is well documented. Although patient-related outcome measures are highly recommended for monitoring overactive bladder symptoms, no study before has dealt with the question of patient-reported complete continence after BoNTA treatment using validated questionnaires.

Clean intermittent self-catheterization after botulinum neurotoxin type A injections: short-term effect on quality of life

OBJECTIVE: To investigate the hypothesis that the need for clean intermittent self-catheterization after botulinum neurotoxin type A injections is outweighed by the efficacy of this treatment, so that clean intermittent self-catheterization is not a burden for patients with refractory idiopathic detrusor overactivity. METHODS: Women undergoing intradetrusor injections of 200 units botulinum neurotoxin type A for refractory idiopathic detrusor overactivity were evaluated prospectively. Clean intermittent self-catheterization was discussed with all patients and its possible need after botulinum neurotoxin type A treatment. As indicator of quality of life, lower urinary tract symptom distress and effect on daily activities were assessed using the validated Urogenital Distress Inventory (UDI-6) and Incontinence Impact Questionnaire (IIQ-7) before and 4 weeks after receiving botulinum neurotoxin type A injections. RESULTS: Mean age of the 65 women was 51 years, and all voided spontaneously before botulinum neurotoxin type A injections. After botulinum neurotoxin type A treatment, 28 (43%) required clean intermittent self-catheterization. Mean UDI-6 and IIQ-7 scores reduced from 61 to 33 (P<.001) and 62 to 30 (P<.001) in women performing clean intermittent self-catheterization and from 60 to 28 (P<.001) and 64 to 25 (P<.001) in those who did not, respectively. Comparison of quality of life in women performing clean intermittent self-catheterization and in those who did not revealed no significant differences before and after botulinum neurotoxin type A treatment. CONCLUSION: Clean intermittent self-catheterization after botulinum neurotoxin type A intradetrusor injections did not impair quality of life in appropriately informed and selected women in the short term. All patients should be informed of the potential need for clean intermittent self-catheterization after botulinum neurotoxin type A injections, and a willingness to do so should be a prerequisite for this still unlicensed off-label treatment.

What a patient with refractory idiopathic detrusor overactivity should know about botulinum neurotoxin type a injection

PURPOSE: We documented the effects of intradetrusor injections of botulinum neurotoxin type A (Botox(R)) for refractory idiopathic detrusor overactivity so that prospective patients maybe properly informed about possible improvement in quality of life, the duration of interinjection intervals and the risk of clean intermittent self-catheterization. MATERIALS AND METHODS: A total of 81 consecutive patients with refractory idiopathic detrusor overactivity treated with intradetrusor injections of 200 U botulinum neurotoxin type A at 20 sites per injection course were evaluated in this prospective, nonrandomized, open label cohort study. The primary outcome was changes in quality of life, as assessed by the short form of the Urogenital Distress Inventory and the Incontinence Impact Questionnaire before and after treatment. Secondary outcomes were the interinjection interval and the need for clean intermittent self-catheterization. RESULTS: After intradetrusor botulinum neurotoxin type A injections there was significant improvement in quality of life, which was sustained after repeat injections. Mean Urogenital Distress Inventory and Incontinence Impact Questionnaire scores decreased from 56 to 26 and 59 to 21 after injection 1 in 81 patients, from 52 to 30 and 51 to 24 after injection 2 in 24, from 40 to 19 and 43 to 17 after injection 3 in 13, from 44 to 17 and 61 to 15 after injection 4 in 6 and from 51 to 17 and 63 to 14 after injection 5 in 4, respectively. The median interinjection interval was 15, 12, 14 and 13 months between injections 1 and 2, 2 and 3, 3 and 4, and 4 and 5, respectively. Considering a post-void residual urine of greater than 100 ml with lower urinary tract symptoms as the indication for clean intermittent self-catheterization, the overall clean intermittent self-catheterization rate after treatment was 43%. CONCLUSIONS: Intradetrusor botulinum neurotoxin type A injections for refractory idiopathic detrusor overactivity significantly improved quality of life. This effect was sustained after repeat injection. More than 2 of 5 patients with refractory idiopathic detrusor overactivity required clean intermittent self-catheterization after botulinum neurotoxin type A injections and all prospective patients should be informed about this.

Suburothelial myofibroblasts in the human overactive bladder and the effect of botulinum neurotoxin type A treatment

BACKGROUND: An increasing body of evidence suggests a possible role of suburothelial myofibroblasts (MFs) in bladder mechanosensation and in the pathophysiology of detrusor overactivity (DO). OBJECTIVE: To determine whether markers of MFs, including gap junction protein connexin43 (Cx43) and c-kit have altered immunohistochemical expression in the suburothelium of patients with neurogenic DO (NDO) or idiopathic DO (IDO) and whether this is affected by successful treatment of DO with botulinum neurotoxin type A (BoNTA). DESIGN, SETTING, AND PARTICIPANTS: Patients with NDO (n=10) or IDO (n=11) were treated in a single-centre, open-label study of intradetrusor BoNTA injections. Control tissue was obtained from 10 patients undergoing pelvic-floor repair procedures who had no overactive bladder (OAB) symptoms. This study is registered with ClinicalTrials.gov, number NCT00662064. INTERVENTIONS: Bladder biopsies performed with flexible cystoscopes were obtained from control subjects and from NDO and IDO patients before BoNTA treatment and at 4 wk and 16 wk after treatment. They were studied with quantitative immunofluorescence using antibodies to connexin 43 (Cx43), vimentin, and c-kit. MEASUREMENTS: Differences in Cx43, vimentin, and c-kit immunoreactivity between control subjects and NDO or IDO patients (primary outcomes). Changes in NDO or IDO, Cx43 immunoreactivity, and c-kit immunoreactivity after BoNTA treatment (secondary outcomes). RESULTS AND LIMITATIONS: Cx43 immunoreactivity was increased in both IDO and NDO patients compared to controls, but remained unchanged after BoNTA treatment. C-kit immunoreactivity was similar in NDO/IDO patients and controls and remained unchanged after BoNTA treatment. CONCLUSIONS: Increased gap junction formation in the suburothelium has been demonstrated in biopsies from humans with DO. It is hypothesised that this change could have a significant role in the pathogenesis of the detrusor abnormality. Successful treatment of NDO or IDO does not appear to be associated with changes in the expression of Cx43 or c-kit on suburothelial MFs.

Immune response of horses to vaccination with the recombinant Hc domain of botulinum neurotoxin types C and D

Botulinum neurotoxins, predominantly serotypes C and D, cause equine botulism through forage poisoning. The C-terminal part of the heavy chain of botulinum neurotoxin types C and D (HcBoNT/C and D) was expressed in Escherichia coli and evaluated as a recombinant mono- and bivalent vaccine in twelve horses in comparison to a commercially available toxoid vaccine. A three-dose subcutaneous immunization of adult horses elicited robust serum antibody response in an ELISA using the immunogen as a capture antigen. Immune sera showed dose-dependent high potency in neutralizing specifically the active BoNT/C and D in the mouse protection assay. The aluminium hydroxide based mono- and bivalent recombinant HcBoNT/C and D vaccines were characterized by good compatibility and the ability to elicit protective antibody titers similar or superior to the commercially available toxoid vaccine.

Alternative vaccination against equine botulism (BoNT/C)

REASON FOR PERFORMING STUDY: In Europe the incidence of botulism in horses has increased in the last decade due to the growing popularity of haylage feeding. Recombinant vaccines are safer and less expensive to produce and are generally better tolerated than toxoids. OBJECTIVES: To investigate whether the recombinant C-terminal half of the heavy chain of the botulinum neurotoxin C (Hc BoNT/C) in combination with an immunstimulatory adjuvant is an appropriate vaccine candidate for horses by testing its efficacy to induce neutralising antibodies and by comparing its immunogenic properties and adverse reactions to a commercial toxoid vaccine. Formation of oedema and local pain reactions were assessed. ELISA and Western blot assay against Hc BoNT/C and testing of neutralising antibody induction in a mouse protection assay were used to evaluate the immune response. RESULTS: With the recombinant vaccine, only minor local swelling with full recovery after 5 days was noted after brisket injections. The toxoid vaccine produced local, painful reactions with longer recovery periods of up to 2 weeks. Horses vaccinated with either vaccine induced neutralising antibodies after the second booster vaccination, while seroconversion on ELISA and Western blot to Hc BoNT/C was apparent after the first recombinant vaccination, and at various time points in the vaccination schedule in horses that received commercial toxoid vaccine. CONCLUSION: The recombinant vaccine showed fewer adverse reactions compared to the only commercially available vaccine but induced similar concentrations of neutralising antibodies. There was no correlation between the serological response to Hc BoNT/C and the neutralising capacity of serum. POTENTIAL RELEVANCE: Recombinant Hc BoNT/C is an appropriate vaccine candidate to stimulate production of neutralising antibodies against botulinum neurotoxin C in horses and creates only minor local reactions at the injection site.

The updated European Consensus 2009 on the use of Botulinum toxin for children with cerebral palsy

An interdisciplinary European group of clinical experts in the field of movement disorders and experienced Botulinum toxin users has updated the consensus for the use of Botulinum toxin in the treatment of children with cerebral palsy (CP). A problem-orientated approach was used focussing on both published and practice-based evidence. In part I of the consensus the authors have tabulated the supporting evidence to produce a concise but comprehensive information base, pooling data and experience from 36 institutions in 9 European countries which involves more than 10,000 patients and over 45,000 treatment sessions during a period of more than 280 treatment years. In part II of the consensus the Gross Motor Function Measure (GMFM) and Gross Motor Function Classification System (GMFCS) based Motor Development Curves have been expanded to provide a graphical framework on how to treat the motor disorders in children with CP. This graph is named "CP(Graph) Treatment Modalities - Gross Motor Function" and is intended to facilitate communication between parents, therapists and medical doctors concerning (1) achievable motor function, (2) realistic goal-setting and (3) treatment perspectives for children with CP. The updated European consensus 2009 summarises the current understanding regarding an integrated, multidisciplinary treatment approach using Botulinum toxin for the treatment of children with CP.

Kinematic improvement following Botulinum Toxin-A injection in upper-limb spasticity due to stroke

Background Focal spasticity is a significant motor disorder following stroke, and Botulinum Toxin Type-A (BoNT-A) is a useful treatment for this. The authors evaluated kinematic modifications induced by spasticity, and whether or not there is any improvement following injection of BoNT-A. Methods Eight patients with stroke with upper-limb spasticity, showing a flexor pattern, were evaluated using kinematics before and after focal treatment with BoNT-A. A group of sex- and age-matched normal volunteers acted as a control group. Results Repeated-measures ANOVA showed that patients with stroke performed more slowly than the control group. Following treatment with BoNT-A, there was a significant improvement in kinematics in patients with stroke, while in the control group, performance remained unchanged. Conclusions Focal treatment of spasticity with BoNT-A leads to an adaptive change in the upper limb of patients with spastic stroke.

Botulinum Toxin Type A for the Treatment of Equinus Deformity in to Patients With Mucopolysaccharidosis Type II

Mucopolysaccharidoses are lysosomal storage disorders that are caused by a deficiency in the enzymes that degrade glycosaminoglycans. The accumulation of glycosaminoglycans affects multiple systems, resulting in coarse facial features, short stature, organomegaly, and variable neurological changes from normal intelligence to severe mental retardation and spasticity. Effects on the musculoskeletal system include dysostosis multiplex, joint stiffness, and muscle shortening. This article reports 2 patients with mucopolysaccharidosis type II (Hunter syndrome) who showed progressive equinus deformity of the feet. Both patients were treated with intramuscular botulinum toxin type A injections in the gastrocnemius and the soleus muscles, followed by serial casting. In both patients, passive range of motion, muscle tone, and gait performance were significantly improved. Botulinum toxin type A injections followed by serial casting are a therapeutic option for contractures in patients with mucopolysaccharidosis. However, the long-term effects and the effect of application in other muscles remain unknown.

A venom-derived neurotoxin, CsTx-1, from the spider Cupiennius salei exhibits cytolytic activities

CsTx-1, the main neurotoxic acting peptide in the venom of the spider Cupiennius salei, is composed of 74 amino acid residues, exhibits an inhibitory cysteine knot motif, and is further characterized by its highly cationic charged C terminus. Venom gland cDNA library analysis predicted a prepropeptide structure for CsTx-1 precursor. In the presence of trifluoroethanol, CsTx-1 and the long C-terminal part alone (CT1-long; Gly-45-Lys-74) exhibit an α-helical structure, as determined by CD measurements. CsTx-1 and CT1-long are insecticidal toward Drosophila flies and destroys Escherichia coli SBS 363 cells. CsTx-1 causes a stable and irreversible depolarization of insect larvae muscle cells and frog neuromuscular preparations, which seem to be receptor-independent. Furthermore, this membranolytic activity could be measured for Xenopus oocytes, in which CsTx-1 and CT1-long increase ion permeability non-specifically. These results support our assumption that the membranolytic activities of CsTx-1 are caused by its C-terminal tail, CT1-long. Together, CsTx-1 exhibits two different functions; as a neurotoxin it inhibits L-type Ca(2+) channels, and as a membranolytic peptide it destroys a variety of prokaryotic and eukaryotic cell membranes. Such a dualism is discussed as an important new mechanism for the evolution of spider venomous peptides.

Comparative aromatic hydroxylation and N-demethylation of MPTP neurotoxin and its analogs, N-methylated beta-carboline and isoquinoline alkaloids, by human cytochrome P450 2D6

1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) neurotoxin is a chemical inducer of Parkinson's disease (PD) whereas N-methylated beta-carbolines and isoquinolines are naturally occurring analogues of MPTP involved in PD. This research has studied the oxidation of MPTP by human CYP2D6 (CYP2D6*1 and CYP2D6*10 allelic variants) as well as by a mixture of cytochrome P450s-resembling HLM, and the products generated compared with those afforded by human monoamine oxidase (MAO-B). MPTP was efficiently oxidized by CYP2D6 to two main products: MPTP-OH (p-hydroxylation) and PTP (N-demethylation), with turnover numbers of 10.09 min-1 and Km of 79.36+/-3 microM (formation of MPTP-OH) and 18.95 min-1 and Km 69.6+/-2.2 microM (PTP). Small amounts of dehydrogenated toxins MPDP+ and MPP+ were also detected. CYP2D6 competed with MAO-B for the oxidation of MPTP. MPTP oxidation by MAO-B to MPDP+ and MPP+ toxins (bioactivation) was up to 3-fold higher than CYP2D6 detoxification to PTP and MPTP-OH. Several N-methylated beta-carbolines and isoquinolines were screened for N-demethylation (detoxification) that was not significantly catalyzed by CYP2D6 or the P450s mixture. In contrast, various beta-carbolines were efficiently hydroxylated to hydroxy-beta-carbolines by CYP2D6. Thus, N(2)-methyl-1,2,3,4-tetrahydro-beta-carboline (a close MPTP analog) was highly hydroxylated to 6-hydroxy-N(2)-methyl-1,2,3,4-tetrahydro-beta-carboline and a corresponding 7-hydroxy-derivative. Thus, CYP2D6 could participate in the bioactivation and/or detoxification of these neuroactive compounds by an active hydroxylation pathway. The CYP2D6*1 enzymatic variant exhibited much higher metabolism of both MPTP and N(2)-methyl-1,2,3,4-tetrahydro-beta-carboline than the CYP2D6*10 variant, highlighting the importance of CYP2D6 polymorphism in the oxidation of these toxins. Altogether, these results suggest that CYP2D6 can play an important role in the metabolic outcome of both MPTP and beta-carbolines.

Botulinum toxin A and B raise blood flow and increase survival of critically ischemic skin flaps

BACKGROUND Botulinum toxin (BTX) A and B are commonly used for aesthetic indications and in neuromuscular disorders. New concepts seek to prove efficacy of BTX for critical tissue perfusion. Our aim was to evaluate BTX A and B in a mouse model of critical flap ischemia for preoperative and intraoperative application. METHODS BTX A and B were applied on the vascular pedicle of an axial pattern flap in mice preoperatively or intraoperatively. Blood flow, tissue oxygenation, tissue metabolism, flap necrosis rate, apoptosis assay, and RhoA and eNOS expression were endpoints. RESULTS Blood-flow measurements 1 d after the flap operation revealed a significant reduction to 53% in the control group, while flow was maintained or increased in all BTX groups (103%-129%). Over 5 d all BTX groups showed significant increase in blood flow to 166-187% (P < 0.01). Microdialysis revealed an increase of glucose and reduced lactate/pyruvate ratio and glycerol levels in the flap tissue of all BTX groups. This resulted in significantly improved tissue survival in all BTX groups compared with the control group (62% ± 10%; all P < 0.01): BTX A preconditioning (84% ± 5%), BTX A application intraoperatively (88% ± 4%), BTX B preconditioning (91% ± 4%), and intraoperative BTX B treatment (92% ± 5%). This was confirmed by TUNEL assay. Immunofluorescence demonstrated RhoA and eNOS expression in BTX groups. All BTX applications were similarly effective, despite pharmacologic dissimilarities and different timing. CONCLUSIONS In conclusion, we were able to show on a vascular, tissue, cell, and molecular level that BTX injection to the feeding arteries supports flap survival through ameliorated blood flow and oxygen delivery.

Equine botulism and acute pasture myodystrophy: new soil-borne emerging diseases in Switzerland?

In Switzerland, the incidence of equine botulism and acute pasture myodystrophy have remarkably increased in the last five years. Equine fodder-borne botulism in Europe is most likely caused by Clostridium botulinum types C and D that produce the toxins BoNT/C and BoNT/D. Horses showing signs suggestive of botulism (muscle weakness and tremors, reduced tongue tone, slow chewing, salivation and difficulties swallowing, drooping eyelids, mydriasis), especially patients that have fed on suspect fodder (mostly haylage), must be treated with anti-serum as soon as possible.They also need intensive care, which is often difficult to provide and always expensive in the face of a guarded to poor prognosis. Therefore, prevention (high standards of forage quality and vaccination) is all the more important. Pasture myodystrophy is an acute disease with signs of rhabdomyolysis and lethality rate over 90%. It affects grazing horses under frosty, windy and rainy conditions. Preliminary results indicate that Clostridium sordellii and Clostridium bifermentans producing lethal toxin may play a role in pasture myodystrophy. Our efforts concentrate on developing a new subunit vaccine for equine botulism and understanding the ethiology and pathogenesis of pasture myodystrophy with the goal of improving prevention against these highly fatal diseases that present a significant risk to our horse population.

Dystonic movement disorders and spinal degenerative disease

The occurrence of degenerative spinal disease subsequent to dystonic movement disorders has been neglected and has received more attention only recently. Spinal surgery is challenging with regard to continuous mechanical stress when treatment of the underlying movement disorder is insufficient. To characterize better the particular features of degenerative spinal disease in patients with dystonia and to analyze operative strategies, we reviewed the available published data. Epidemiologic studies reveal that degenerative spinal disorders in patients with dystonia and choreoathetosis occur much earlier than in the physiological aging process. Dystonic movement disorders more often affect the spine at higher cervical levels (C(2-5)), in contrast to spinal degeneration with age which manifests more frequently at the middle and lower cervical spine (C(5-7)). Degenerative changes of the cervical spine are more likely to occur on the side where the chin is rotated or tilted to. Various operative approaches for treatment of spinal pathologies have been advocated in patients with dystonic movement disorders. The available data do not allow making firm statements regarding the superiority of one approach over the other. Posterior approaches were first used for decompression, but additional anterior fusion became necessary in many instances. Anterior approaches with or without instrumented fusion yielded more favorable results, but drawbacks are pseudarthrosis and adjacent-level disease. Parallel to the development of posterior fusion techniques, circumferential surgery was suggested to provide a maximum degree of cord decompression and a higher fusion rate. Perioperative local injections of botulinum toxin were used initially to enhance patient comfort with halo immobilization, but they are also applied in patients without external fixation nowadays. Treatment algorithms directed at the underlying movement disorder itself, taking advantage of new techniques of functional neurosurgery, combined with spinal surgery have recently been introduced and show promising results.