Continuous evaluation of single-dose moxifloxacin concentrations in brain extracellular fluid, cerebrospinal fluid, and plasma: a novel porcine model.

BACKGROUND: Knowledge regarding CNS pharmacokinetics of moxifloxacin is limited, with unknown consequences for patients with meningitis caused by bacteria resistant to beta-lactams or caused by TB. OBJECTIVE: (i) To develop a novel porcine model for continuous investigation of moxifloxacin concentrations within brain extracellular fluid (ECF), CSF and plasma using microdialysis, and (ii) to compare these findings to the pharmacokinetic/pharmacodynamic (PK/PD) target against TB. METHODS: Six female pigs received an intravenous single dose of moxifloxacin (6 mg/kg) similar to the current oral treatment against TB. Subsequently, moxifloxacin concentrations were determined by microdialysis within five compartments: brain ECF (cortical and subcortical) and CSF (ventricular, cisternal and lumbar) for the following 8 hours. Data were compared to simultaneously obtained plasma samples. Chemical analysis was performed by high pressure liquid chromatography with mass spectrometry. The applied PK/PD target was defined as a maximum drug concentration (Cmax):MIC ratio >8. RESULTS: We present a novel porcine model for continuous in vivo CNS pharmacokinetics for moxifloxacin. Cmax and AUC0-8h within brain ECF were significantly lower compared to plasma and lumbar CSF, but insignificantly different compared to ventricular and cisternal CSF. Unbound Cmax:MIC ratio across all investigated compartments ranged from 1.9 to 4.3. CONCLUSION: A single dose of weight-adjusted moxifloxacin administered intravenously did not achieve adequate target site concentrations within the uninflamed porcine brain ECF and CSF to reach the applied TB CNS target.

Analysis of free cisplatin in microdialysates and plasma ultrafiltrate by liquid chromatography-tandem mass spectrometry.

Cisplatin is a potent cytotoxic agent used in the treatment of various malignancies and exerts its antitumor effect through malignant cell DNA damage and apoptosis induction. Evaluation of systemic delivery of cisplatin is important in optimization of cisplatin treatment. However, accurate quantification of systemic cisplatin is challenging due to its various forms in circulation. This study aimed to develop a sensitive (LOQ < 0.1 µg/mL) and precise Ultra Performance Liquid Chromatography (UPLC) - Tandem Mass Spectrometry (MS/MS) method for quantifying free cisplatin in microdialysates and plasma. Furthermore the aim was to compare free cisplatin concentrations measured in standard plasma samples with those obtained from intravenous microdialysis catheters in a porcine model. The method developed utilizes dichloro(ethylenediamine)platinum(II) as an internal standard that co-elutes with cisplatin, ensuring precise correction for ion suppression/enhancement effects. The method was validated, demonstrating linearity up to 100 µg/mL and good intermediate precision (CV% < 6 %) in the range of 1.0-100 µg/mL, with an LOQ of 0.03 µg/mL. The pharmacokinetic parameters (AUC0-last, Cmax, T1/2, and Tmax) showed no significant differences between the two sampling methods. This validated LC-MS/MS method provides a reliable tool for quantifying systemic free cisplatin concentrations, facilitating future systemic and local pharmacokinetic evaluations for optimization of cisplatin-based cancer treatments.

Double-dose cefuroxime concentrations in bone, synovial fluid of the knee joint and subcutaneous adipose tissue-A randomised porcine microdialysis study.

This study evaluated target tissue concentrations of double dose cefuroxime administered intravenously as either one 15 min infusion of 3000 mg (Group 1) or two single 15 min infusions of 1500 mg administered 4 h apart (Group 2). Sixteen pigs were randomised into two groups of eight. Cortical and cancellous bone, synovial fluid of the knee joint and subcutaneous adipose tissue concentrations were measured based on sampling via microdialysis. Plasma samples were collected as a reference. Comparison of the groups was based on time with concentrations above relevant minimal inhibitory concentrations (fT>MIC) of 4 µg/mL. The mean time fT>MIC (4 µg/mL) across compartments was longer for Group 2 (280-394 min) than for Group 1 (207-253 min) (p<0.01). Cortical bone showed a tendency towards longer fT>MIC (4 µg/mL) in Group 2 (280 min) than in Group 1 (207 min) (p = 0.053). Within 50 min after administration, the mean concentration of 4 µg/mL was reached in all compartments for both groups. The mean concentrations decreased below 4 µg/mL after approximately 4 h (Group 1) and 3 h (Group 2) from initiation of administration (time zero). During an 8 h interval, double-dose cefuroxime administered as 2 × 1500 mg with a 4 h interval provides longer time above MIC breakpoint for Staphylococcus aureus (4 µg/mL) than a single bolus of 3000 mg cefuroxime. To maintain sufficient tissue concentrations during longer surgeries, re-administration of cefuroxime (1500 mg) should be considered 3 h after the first administration.

Allogeneic adipose tissue-derived mesenchymal stem cells in combination with platelet rich plasma are safe and effective in the therapy of superficial digital flexor tendonitis in the horse.

Overstrain tendonitis are common pathologies in the sport horses. Therapeutic approaches to tendon healing do not always result in a satisfactory anatomical and functional repair, and healed tendon is often characterized by functional impairment and high risk of reinjury. Recently, mesenchymal stem cells (MSCs) and platelet rich plasma (PRP) have been proposed as novel therapeutic treatments to improve the tendon repair process. MSCs are multipotent, easy to culture and being originated from adult donors do not pose ethical issues. To date, autologous MSCs have been investigated mainly in the treatment of large bone defects, cardiovascular diseases, osteogenesis imperfecta and orthopaedic injuries both in human and veterinary medicine. The clinical applications in which autologous MSCs can be used are limited because patient-specific tissue collection and cell expansion require time. For clinical applications in which MSCs should be used right away, it would be more practical to use cells collected from a donor, expanded in vitro and banked to be readily available when needed. However, there are concerns over the safety and the efficacy of allogeneic MSCs. The safety and efficacy of a therapy based on the use of allogeneic adipose tissue-derived mesenchymal stem cells (ASCs) associated to platelet rich plasma (PRP) were evaluated in 19 horses affected by acute or subacute overstrain superficial digital flexor tendonitis (SDFT). The application of allogeneic ASCs neither raised clinical sign of acute or chronic adverse tissue reactions, nor the formation of abnormal tissue in the long-term. After a follow-up of 24 months, 89.5% horses returned to their previous level of competition, while the reinjury rate was 10.5%, comparable to those recently reported for SDFT treated with autologous bone marrow derived MSCs. This study suggests that the association between allogeneic ASCs and PRP can be considered a safe and effective strategy for the treatment of SDF tendonitis in the horse.

[Clinical study of a cohort of 94 children with influenza A(H1N1) on the Réunion Island]. / Étude clinique d'une cohorte de 94 enfants atteints de la grippe A(H1N1)v à l'île de La Réunion.

We conducted a monocentric retrospective study at the Félix-Guyon Regional Hospital, Saint-Denis, Réunion Island, from August 1, 2009 to September 30, 2009. We included 94 children with nasal and/or throat swabs tested positive for influenza A(H1N1)v during the study period. We describe the clinical, biological, and radiological presentations, the hospitalization rate, and the antiviral and antibiotic therapies. Children between six months and two years were the ones most at risk of severe forms and the first risk factor was asthma. Children with risk factors were at risk of severe forms except immune suppressed children. 7% of our cohort had only digestive signs. Recommendations for antiviral treatment were followed in 75% of the treated children. Risk factors and clinical signs, not age, guided physicians in the initiation of treatment.

[Alcohol and pregnancy]. / Alcool et grossesse.

Alcohol consumption during pregnancy is a major cause of mental retardation in Western countries. Fetal alcohol syndrome (FAS) is mainly characterized by pre- and postnatal stunted growth, neurocognitive disorders, and facial dysmorphism. It compromises the intellectual and behavioral prognosis of the child. Prevention tools exist, through better information of health professionals, for optimal care of high-risk women before, during, and after pregnancy, which would decrease the incidence of SAF in the future.

Measurement of magnetic losses by thermal method applied to power ferrites at high level of induction and frequency.

Classically, low frequency losses in soft magnetic materials and ferrites in particular are measured by flux metric method under sinusoidal waveform excitation voltage. However, in typical application of modern power electronics, the frequency currently exceeds 100 kHz. This feature is at the origin of a difficulty: the phase shift between current and voltage can be disturbed by current probe delay. Thus, the results can be affected by large errors. As a consequence, it becomes more and more important to develop alternative methods to measure losses in magnetic materials. It is proposed to use calorimetric method which is by principle free of the above mentioned problems. The experimental device is described in details and the results are reported for experiments conducted on a commercial Mn-Zn ferrite under sinusoidal waveform regime for frequencies varying from 10 to 200 kHz. Comparisons with flux metric measurement show that significant differences appear typically for Bf products above 5000 V/m(2) (50 kHzx100 mT).

Equine adipose-tissue derived mesenchymal stem cells and platelet concentrates: their association in vitro and in vivo.

Equine mesenchymal stem cells (MSC) are of particular interest both for basic research and for the therapeutic approach to musculoskeletal diseases in the horse. Their multilineage differentiation potential gives them the capability to contribute to the repair of tendon, ligament and bone damage. MSCs are also considered a promising therapeutic aid in allogeneic cell transplantation, since they show low immunogenicity and immunomodulating functions.Adipose tissue-derived adult equine stem cells (AdMSC) can be isolated, expanded in vitro and then inoculated into the damaged tissue, eventually in the presence of a biological scaffold. Here we report our preliminary experience with adipose-derived mesenchymal stem cells in allogeneic cell-therapy of tendonitis in the horse. MSCs, derived from visceral adipose tissue, were grown in the presence of autologous platelet lysate and characterized for their differentiation and growth potential. Expanded AdMSC were inoculated into the damaged tendon after their dispersion in activated platelet-rich plasma (PRP), a biological scaffold that plays an important role in maintaining cells in defect sites and contributes to tissue healing. Fourteen out of sixteen treated horses showed a functional recovery and were able to return to their normal activity.

Focal canine tetanus: diagnostic value of electromyography.

A four-year-old, male Italian hound was presented with severe spasticity of both thoracic limbs that worsened with external stimuli. The remainder of the neurological and general physical examination was normal. Complete blood cell count, chemistry profile, and serology for protozoal diseases were within normal limits. Survey radiography of the cervicothoracic spine and abdominal ultrasonography showed no abnormalities. Electromyography of the thoracic limbs demonstrated the presence of "doublets" and simultaneous activity in both agonist and antagonist muscles. These abnormalities may be explained by a defective glycinergic inhibition at the spinal cord level. Together with the history, progression of signs, and clinical findings, electromyography supported a presumptive diagnosis of focal tetanus. The dog received tetanus antitoxin and antibiotic treatment and gradually improved over four months.

Incomplete humeral condylar fracture in two English Pointer dogs.

Incomplete humeral condylar fracture was diagnosed by means of radiology, CT, scintigraphy, arthroscopy and bone biopsy in two English Pointer dogs. In both cases an acute thoracic limb lameness, unrelated to a known episode of major trauma, was observed. Incomplete humeral condylar fracture, mainly described in the Spaniel breeds, has been recently diagnosed in Labrador retrievers, Rottweiler, German Wachtel and other breeds. The pathogenesis of the condition is still unknown, but incomplete ossification of the humeral condyle and mechanical stress, alone or associated, have to be considered. However, our clinical and histopathological data lead us to believe that in Pointers, high performance dogs, the mechanical stress can assume a critical ethiopathogenetic role.

Cyclin D1 expression in pre-cancerous and cancerous lesions of the canine mammary gland.

Overexpression of cyclin D1, the regulatory subunit of cyclin-dependent kinases (cdk4 and cdk6) involved in cell cycle control, has often been found in breast cancer and other types of human cancer. Increased expression, or stability, of cyclin D1 molecules may cause sufficient cdk4 activation to produce retinoblastoma protein phosphorylation independently of mitogenic signals; this results in commitment of cells to the G1 phase at mitosis. In the present study, cyclin D1 expression was investigated in pre-cancerous and cancerous lesions of the canine mammary gland by a complex experimental approach, which included Western blot and immunohistochemical analysis of cyclin D1 and the related molecular system. Furthermore, to define relationships between cell growth and expression of cyclin D1, proliferative activity was studied by the AgNOR technique. The study provided the following information. Cyclin D1 overexpression was largely independent of the type of proliferative anomaly. Indeed, cyclin D1 was expressed in 60% of the pre-cancerous lesions and in 44% of cancerous lesions. Mitotic activity and cyclin D1 expression were related: mammary lesions that expressed cyclin D1 showed a high proliferative ratio, the opposite being true of cyclin D1-negative cell populations. This study may contribute to the establishment of an animal model for anti-cancer research based on cyclin D1 suppression or cdk inactivation, or both.

A simplified technique for diagnostic and surgical arthroscopy of the shoulder joint in the dog.

A modified technique is presented for surgical and diagnostic arthroscopy of the shoulder joint in the dog. The technique involves access to the joint through two points only; one was created in place of the drainage needle-cannula, which was replaced with a portal, while the second was located more caudally compared with previous techniques. Using a changing guide rod system the two portals are completely interchangeable in order to perform easier arthroscopic surgery either in the cranial or caudal aspect of the joint. The presence of only one portal caudal to the lateral collateral ligament allows more freedom of movement and avoids interference between the arthroscope and the instruments. The modified procedure was performed on 33 joints affected by osteochondritis dissecans or tenoligament diseases and facilitated straightforward diagnostic examinations, and simple and rapid surgical procedures.

Extra-articular absorbable suture stabilization of coxofemoral luxation in dogs.

OBJECTIVE: To evaluate the effectiveness of an extra-articular surgical technique using absorbable suture material for the stabilization of traumatic coxofemoral luxation in dogs. STUDY DESIGN: Prospective, clinical study. ANIMALS: Fourteen client-owned dogs with recent and long-standing traumatic coxofemoral luxation (13 craniodorsal and 1 ventral). METHODS: Coxofemoral luxations were surgically reduced and maintained in place with an extra-articular iliofemoral multifilamentous absorbable suture (3 to 6 strands of 2 USP Polyglactin 910). No external support was employed, and all the dogs were encouraged to use the affected limb postoperatively. The average time of clinical and radiographic follow-up was 11.6 +/- 6.3 months (from 2 to 22 months). RESULTS: During the follow-up period, no reluxations occurred and no complications associated with the surgical technique were identified. The dogs started bearing weight from 1 to 10 days after the surgery (mean, 4.3 +/- 2.9 days) and the period of lameness ranged from 7 to 30 days (20 +/- 8.6 days). At the final clinical examination, the dogs did not demonstrate any lameness or pain during passive flexo-extension movements, and there was no significant limitation of the range of motion. CONCLUSION: Extra-articular stabilization with multifilamentous absorbable sutures is a simple, effective method of treatment for acute and chronic coxofemoral luxation. The absorbable material used is strong enough to maintain articular stability during the period of scar tissue formation even in large-breed dogs. CLINICAL RELEVANCE: Absorbable sutures avoid the possible complications related to the use of nonabsorbable material and seem to be sufficient to maintain articular stability during the capsular healing process.

The inhibitory effects of interleukin-6 on synaptic plasticity in the rat hippocampus are associated with an inhibition of mitogen-activated protein kinase ERK.

Several cytokines have short-term effects on synaptic transmission and plasticity that are thought to be mediated by the activation of intracellular protein kinases. We have studied the effects of interleukin-6 (IL-6) on the expression of paired pulse facilitation (PPF), posttetanic potentiation (PTP), and long-term potentiation (LTP) in the CA1 region of the hippocampus as well as on the activation of the signal transducer and activator of transcription-3 (STAT3), the mitogen-activated protein kinase ERK (MAPK/ERK), and the stress-activated protein kinase/c-Jun NH(2)-terminal kinase (SAPK/JNK). IL-6 induced a marked and dose-dependent decrease in the expression of PTP and LTP that could be counteracted by the simultaneous treatment with the tyrosine kinase inhibitor lavendustin A (LavA) but did not significantly affect PPF. The IL-6-induced inhibition of PTP and LTP was accompanied by a simulation of STAT3 tyrosine phosphorylation and an inhibition of MAPK/ERK dual phosphorylation, in the absence of changes in the state of activation of SAPK/JNK. Both effects of IL-6 on STAT3 and MAPK/ERK activation were effectively counteracted by LavA treatment. The results indicate the tyrosine kinases and MAPK/ERK are involved in hippocampal synaptic plasticity and may represent preferential intracellular targets for the actions of IL-6 in the adult nervous system.

Loss of GSH, oxidative stress, and decrease of intracellular pH as sequential steps in viral infection.

Madin-Darby canine kidney cells infected with Sendai virus rapidly lose GSH without increase in the oxidized products. The reduced tripeptide was quantitatively recovered in the culture medium of the cells. Since the GSH loss in infected cells was not blocked by methionine, a known inhibitor of hepatocyte GSH transport, a nonspecific leakage through the plasma membrane is proposed. UV-irradiated Sendai virus gave the same results, confirming that the major loss of GSH was due to membrane perturbation upon virus fusion. Consequent to the loss of the tripeptide, an intracellular pH decrease occurred, which was due to a reversible impairment of the Na+/H+ antiporter, the main system responsible for maintaining unaltered pHi in those cells. At the end of the infection period, a rise in both pHi value and GSH content was observed, with a complete recovery in the activity of the antiporter. However, a secondary set up of oxidative stress was observed after 24 h from infection, which is the time necessary for virus budding from cells. In this case, the GSH decrease was partly due to preferential incorporation of the cysteine residue in the viral proteins and partly engaged in mixed disulfides with intracellular proteins. In conclusion, under our conditions of viral infection, oxidative stress is imposed by GSH depletion, occurring in two steps and following direct virus challenge of the cell membrane without the intervention of reactive oxygen species. These results provide a rationale for the reported, and often contradictory, mutual effects of GSH and viral infection.

Glutathione inhibits HIV replication by acting at late stages of the virus life cycle.

We investigated the effect of glutathione on the replication of human immunodeficiency virus (HIV) in chronically infected macrophages, a known reservoir of the virus in the body. We found that exogenous GSH strongly suppresses the production of p24gag protein as well as the virus infectivity. This is related to a dramatic decrease in both budding and release of virus particles from chronically infected cells (either macrophages or lymphocytes), together with a selective decrease in the expression of gp120, the major envelope glycoprotein, rich in intrachain disulfide bonds and thus potentially sensitive to the effect of a reducing agent such as GSH. Overall data suggest that GSH can interfere with late stages of virus replication. This would be in agreement with data obtained in cells exposed to herpesvirus type 1 (a DNA virus) or to Sendai (an RNA virus), showing that the suppression of virus replication by GSH is related to the selective inhibition of envelope glycoproteins. These results suggest a potential role of GSH in combination with other antivirals in the treatment of virus-related diseases.

[Tension-free laparoscopic hernioplasty]. / Ernioplastica "tension free" per via laparoscopica.

Hernia recurrence after traditional "open" hernioplasty is an observed event. The tension undergone by anatomical structures of the area is believed to be responsible in large part for these failures. Thus, techniques involving "tension free" hernioplasty have been developed, some of which involve laparoscopic access. Here an experience is reported regarding laparoscopic hernioplasty carried out to repair groin hernias, first of all on an animal model (pigs: 7 cases) then in the clinical field. The technique chosen was the endo-peritoneal positioning of a PTFE prosthesis. The results revealed several advantages over the traditional methods, basically the possibility of reinforcing the entire inguinal floor at the same time as repairing the hernia, and the decrease in groin area discomfort and less time off from work for the patient.

[Experimental intestinal laparoscopic resection]. / Resezione intestinale laparoscopica sperimentale: note di tecnica.

At present, there is a debate in the literature regarding the possibility of extending the laparoscopic approach to intestinal resection. Some techniques have been developed, but certain imperfections and problems remain. This experimental contribution suggests answers to some technical problems arising in the course of a small-bowel laparoscopic resection on a pig.

Intraarticular sodium hyaluronate injections in the Pond-Nuki experimental model of osteoarthritis in dogs. I. Biochemical results.

An established anterior cruciate ligament deficiency-induced articular cartilage degeneration was used to evaluate the effects of intrasynovial injection of hyaluronic acid upon cartilage destruction. In this study, proteoglycan solubility under associative and dissociative conditions was compared in two treatment protocols at intervals of seven, 13, and 17 weeks after surgical breakage of the anterior cruciate ligament in 2.5-year-old Beagle dogs. Untreated joints showed a marked increase in both total soluble glycosaminoglycan (GAG measured as uronic acid) and in the associative fraction. In both treated groups, there was a reduced amount of soluble GAG. Cessation of treatment after seven weeks caused gradual regression, with an increasing amount of CaCl2-soluble material in the associative fraction, while inception at seven weeks gave biochemical evidence of reversal, with increasing GAG present in the guanidine-soluble (dissociative) fraction on the insoluble residue. The protective effects of hyaluronic acid suggest the potential clinical application of this therapy in retarding the advance of osteoarthritis.