Anthelmintic activity of cyclotides: In vitro studies with canine and human hookworms

Hookworm infection is a leading cause of maternal and child morbidity in countries of the tropics and subtropics, as well as being an important parasite in companion-animal medicine. The cyclotides are a novel family of cyclic cystine knot containing peptides from plants that have been shown to possess anthelmintic activity against Haemonchus contortus and Trichostrongylus colubriformis, two important gastrointestinal nematodes of sheep. In the current study we demonstrated the in vitro effects of three representative cyclotides, kalata B1, kalata B6 and cycloviolacin O14, on the viability of larval and adult life stages of the dog hookworm Ancylostoma caninum, and larvae of the human hookworm Necator americanus. The cyclotides showed significant anthelmintic activity towards both hookworm species. The different cyclotides showed similar patterns of relative activity as that seen previously with the livestock nematode species. This study demonstrates that cyclotides have promising activity in vitro against important parasites of companion animals and humans.     

Postinflammatory ossicular fixation: CT analysis with surgical correlation

Postinflammatory ossicular fixation is a common problem encountered by the otologic surgeon upon exploration because of conductive hearing loss in patients with chronic otitis media. These nonotosclerotic noncongenital lesions take three pathologic forms: fibrous tissue fixation (chronic adhesive otitis media), hyalinization of collagen (tympanosclerosis), and new bone formation (fibro-osseous sclerosis). Fibrous tissue fixation appears on CT as nonbony, noncalcific soft-tissue debris encasing some or all of the ossicular chain. Tympanosclerosis appears as unifocal or multifocal punctate or weblike calcifications in the middle ear cavity or on the tympanic membrane. This debris may be in direct apposition to the ossicular chain or may replace the suspensory ligaments in symptomatic patients. New bone formation has been identified only in the attic and is the least common manifestation. Thick bony webs or generalized bony encasement may be present at CT. More than 300 patients with the clinical diagnosis of chronic otitis media have been examined. This study encompasses 23 proved cases.     

Rats recovering from unilateral barrel-cortex ischemia are capable of completing a whisker-dependent task using only their affected whiskers

Rats use their vibrissae for a variety of exploratory tasks including location of objects and discrimination of texture. This study examines recovery in vibrissal function following a unilateral ischemic injury to the somatosensory cortex. Vibrissal function was examined in adult food-restricted rats performing on a two-texture discrimination device. Animals were trained and tested until the criteria of >80% correct choices was demonstrated on three consecutive days. Ischemic rats were constrained to use the affected whiskers by clipping the ipsilateral vibrissae. One group was tested after ischemia, a second group was trained before ischemia and then tested, and a third group was pre-trained and received whisker stimulation and tested post-ischemia. Nai;ve animals recovering from ischemia took longer to reach criteria than intact or unilateral trimmed control animals. Pre-trained animals with compression ischemia receiving whisker stimulation with sucrose water completed the task to criteria in the fewest number of trials. The results indicate that recovery of vibrissal function occurs following a unilateral ischemic injury. Histological analysis in animals without whisker stimulation indicates that the number of normal appearing cortical barrels following ischemia was inversely correlated to the number of trials needed to complete the behavioral task. This suggests that the natural recovery of the ability to discriminate textures is related to the degree of damage to the barrel cortex. The relationship between cortical barrels and behavioral recovery did not hold for the ischemic animals receiving whisker stimulation. This latter group demonstrated recovery despite marked anatomical lesions suggesting that the intervention influenced reorganization.     

Fat-to-glucose interconversion by hydrodynamic transfer of two glyoxylate cycle enzyme genes

The glyoxylate cycle, which is well characterized in higher plants and some microorganisms but not in vertebrates, is able to bypass the citric acid cycle to achieve fat-to-carbohydrate interconversion. In this context, the hydrodynamic transfer of two glyoxylate cycle enzymes, such as isocytrate lyase (ICL) and malate synthase (MS), could accomplish the shift of using fat for the synthesis of glucose. Therefore, 20 mice weighing 23.37 ± 0.96 g were hydrodinamically gene transferred by administering into the tail vein a bolus with ICL and MS. After 36 hours, body weight, plasma glucose, respiratory quotient and energy expenditure were measured. The respiratory quotient was increased by gene transfer, which suggests that a higher carbohydrate/lipid ratio is oxidized in such animals. This application could help, if adequate protocols are designed, to induce fat utilization for glucose synthesis, which might be eventually useful to reduce body fat depots in situations of obesity and diabetes.     

Chromosomal assignments of human genes for serine proteases trypsin,chymotrypsin B,and elastase

The genes for the serine proteases trypsin, chymotrypsin B, and elastase were chromosomally assigned in man using cDNA probes that have been isolated from a rat pancreatic cDNA library. DNA from human × rodent somatic cell hybrids was cleaved with BamHI or EcoRI and analyzed by Southern filter hybridization methods for the segregation of the genes for trypsin-1 (TRY1), chymotrypsin B (CTRB), and elastase-1 (ELA1). TRY1 was assigned to human chromosome 7q22qter, CTRB to chromosome 16, and ELA1 to chromosome 12. Although the three genes are members of the same gene family, they are dispersed over different chromosomes.Preliminary chromosome assignments of chymotrypsin B (1) and of trypsin and elastase (2) have been made in abstract.     

Chirurgische Behandlung infizierter Knochendefekte

Zusammenfassung Die unhappy triad der Knochenchirurgie, Infekt, Defekt und Instabilität stellt uns auch heute noch vor schwer zu lösende Probleme. Das Vorgehen der Wahl scheint uns die Stabilisierung der Fragmente mittels einer internen (Osteosynthese) oder externen (äußere Spanner) Fixation, die radikale Ausräumung des Herdes, die vorübergehende Spüldrainage nach Willenegger [26, 27, 28] und schließlich das Auffüllen des Defektes mit autologer Spongiosa zu sein. Bei allen unseren in dieser Studie erfaßten 25 Patienten kam es zum knöcherner Einbau des Transplantates und Abheilung des Haut- und Weichteildefektes, bei vier Patienten be schleunigte eine Spalthautverpflanzung die Heilung. 23 Fälle sind 1 bis 6 Jahre nach der Behandlung vom Infekt her rezidivfrei geblieben, bei sämtlichen Patienten konnte die Belastungsstabilität erreicht werden.Durch das beschriebene Vorgehen konnte in allen Fällen die Gelenkfunktion erhalten oder verbessert werden. Das aktive Eingreifen gestattet zudem Achsen- und Längenkorrekturen.Klinische, szintigraphische und histologische Untersuchungen zeigen, daß der Einbau des spongiösen Transplantates unmittelbar nach der Verpflanzung einsetzt und nach 3 Monaten soweit fortgeschritten ist, daß die Belastungsstabilität erreicht wird.

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Autogenous cancellous bone in osteomyelitis with defects of bone, soft tissue and skin

Summary Surgeons are still confronted with the grave problem of the unhappy triad of bone surgery, i.e. infection, osseous defect and instability. To us the stabilisation of fragments by means of internal or external fixation, the radical saucerization and packing of the cavity with autogenous cancellous bone with preceding irrigation drainage seems to be the procedure best suitable. 25 patients with infected defects of bone, soft tissue and skin were treated accordingly and followed up 1 to 6 years later: In all cases the graft had been integrated and the skin- and soft tissue defects had healed. In 23 cases osteomyelitis had not re-occured, weight bearing stability had been achieved in all 25 cases.The described procedure had either maintained or even improved articular function. Additionally the active intervention allowes correction of axis and length.It is demonstrated by radiological, scintigraphical and histological examinations, that the integration of the cancellous bone transplant begins immediately after transplantation and is advanced within three months to such a point that weight bearing becomes possible.

     

Substrates of the prostate-specific serine protease prostase/KLK4 defined by positional-scanning peptide libraries

BACKGROUND: Prostase/KLK4 is a member of the human kallikrein (KLK) gene family that is expressed in prostate epithelial cells under the regulation of androgenic hormones. In this study, we sought to characterize the substrate specificity of KLK4 in order to gain insight into potential physiological roles of the enzyme. METHODS: A chimeric form of KLK4 was constructed in which the pro-region of KLK4 was replaced with the signal and propeptide sequence of trypsinogen (proT-KLK4) to create an activation site susceptible to enterokinase cleavage. proT-KLK4 was expressed in Drosophila S2 cells, purified, and activated with enterokinase to generate mature KLK4. The extended substrate specificity of KLK4 was defined by screening tetrapeptide positional scanning synthetic combinatorial libraries (PS-SCL). RESULTS: The preferred P1-P4 positions as determined by PS-SCL were: P1-Arg; P2-Gln/Leu/Val; P3-Gln/Ser/Val; P4-Ile/Val. The trypsin-like specificity of KLK4 was further confirmed using synthetic chromogenic peptides. Based upon the optimal cleavage site residues, a database search for potential KLK4 substrates identified several proteins with potential roles mediating normal prostate physiology or neoplastic growth including KLK3/PSA, parathyroid hormone-related peptide (PTHrP), and members of the bone morphogenetic protein (BMP) family. Recombinant KLK4 was able to activate pro-PSA/KLK3 and degrade members of the insulin-like growth factor (IGF) binding protein (IGFBP) family. CONCLUSIONS: These results identify potential KLK4 substrates that may serve to define the role of this protease in normal prostate physiology, and facilitate studies of the consequences of KLK4 expression in pathological conditions.     

Task-related activity in prefrontal cortex and its relation to recognition memory performance in young and old adults

Older adults often have more widespread prefrontal cortex (PFC) activation during memory retrieval tasks, compared to young adults, particularly in the left hemisphere. Recruitment of additional frontal activity in older adults has been attributed by some researchers to compensation, perhaps for reduced activity elsewhere in the brain, whereas others have described it as a non-selective response that may be due to a failure to inhibit these PFC regions. To address further the impact of PFC activity on memory in older adults, we used PET to measure brain activity during recognition memory tasks. Both young and old adults showed increased activity during recognition, compared to a control task, in bilateral PFC. Young adults showed greater activation of left hippocampus and lateral temporal cortex during recognition, whereas older adults showed greater activity in the right inferior frontal gyrus. Age differences also were seen in correlations between brain activity and memory performance. There were positive correlations between activity in the right parahippocampal gyrus and recognition performance in young adults, whereas positive correlations between activity in PFC and performance were found only in older adults. These positive correlations included the right inferior PFC region where older adults had greater activation. Activity in this right PFC region was negatively correlated with medial temporal activity in both groups. These results provide further evidence for age-specific patterns of brain activity underlying memory performance and are consistent with the idea that PFC assumes a larger role in supporting successful recognition memory with increasing age. The negative correlation between activity in PFC and medial temporal regions, as well as the age differences in how these regions were related to behavior, suggest that those older individuals who recruit PFC to a greater degree may do so as a compensatory response to reductions in medial temporal regions.     

RegIIA: an α4/7-conotoxin from the venom of Conus regius that potently blocks α3β4 nAChRs

Neuronal nicotinic acetylcholine receptors (nAChRs) play pivotal roles in the central and peripheral nervous systems. They are implicated in disease states such as Parkinson's disease and schizophrenia, as well as addictive processes for nicotine and other drugs of abuse. Modulation of specific nAChRs is essential to understand their role in the CNS. α-Conotoxins, disulfide-constrained peptides isolated from the venom of cone snails, potently inhibit nAChRs. Their selectivity varies markedly depending upon the specific nAChR subtype/α-conotoxin pair under consideration. Thus, α-conotoxins are excellent probes to evaluate the functional roles of nAChRs subtypes. We isolated an α4/7-conotoxin (RegIIA) from the venom of Conus regius. Its sequence was determined by Edman degradation and confirmed by sequencing the cDNA of the protein precursor. RegIIA was synthesized using solid phase methods and native and synthetic RegIIA were functionally tested using two-electrode voltage clamp recording on nAChRs expressed in Xenopus laevis oocytes. RegIIA is among the most potent antagonist of the α3β4 nAChRs found to date and is also active at α3β2 and α7 nAChRs. The 3D structure of RegIIA reveals the typical folding of most α4/7-conotoxins. Thus, while structurally related to other α4/7 conotoxins, RegIIA has an exquisite balance of shape, charge, and polarity exposed in its structure to potently block the α3β4 nAChRs.