New frontiers in applied veterinary point‐of‐capture diagnostics: Toward early detection and control of zoonotic influenza

Among the chief limitations in achieving early detection and control of animal‐origin influenza of pandemic potential in high‐risk livestock populations is the existing lag time between sample collection and diagnostic result. Advances in molecular diagnostics are permitting deployment of affordable, rapid, highly sensitive, and specific point‐of‐capture assays, providing opportunities for targeted surveillance driving containment strategies with potentially compelling returns on investment. Interrupting disease transmission at source holds promise of disrupting cycles of animal‐origin influenza incursion to endemicity and limiting impact on animal production, food security, and public health. Adoption of new point‐of‐capture diagnostics should be undertaken in the context of promoting robust veterinary services systems and parallel support for operationalizing pre‐authorized plans and communication strategies that will ensure that the full potential of these new platforms is realized.     

Plasmafiltration as an effective method in the removal of circulating pegylated liposomal doxorubicin (PLD) and the reduction of mucocutaneous toxicity during the treatment of advanced platinum-resistant ovarian cancer

Cancer Chemotherapy and Pharmacology - The present study evaluates the safety and efficacy of double-plasma filtration (PF) to remove the exceeding pegylated liposomal doxorubicin (PLD) in...     

Motion‐compensated b‐tensor encoding for in vivo cardiac diffusion‐weighted imaging

Motion is a major confound in diffusion‐weighted imaging (DWI) in the body, and it is a common cause of image artefacts. The effects are particularly severe in cardiac applications, due to the nonrigid cyclical deformation of the myocardium. Spin echo‐based DWI commonly employs gradient moment‐nulling techniques to desensitise the acquisition to velocity and acceleration, ie, nulling gradient moments up to the 2nd order (M2‐nulled). However, current M2‐nulled DWI scans are limited to encode diffusion along a single direction at a time. We propose a method for designing b‐tensors of arbitrary shapes, including planar, spherical, prolate and oblate tensors, while nulling gradient moments up to the 2nd order and beyond. The design strategy comprises initialising the diffusion encoding gradients in two encoding blocks about the refocusing pulse, followed by appropriate scaling and rotation, which further enables nulling undesired effects of concomitant gradients. Proof‐of‐concept assessment of in vivo mean diffusivity (MD) was performed using linear and spherical tensor encoding (LTE and STE, respectively) in the hearts of five healthy volunteers. The results of the M2‐nulled STE showed that (a) the sequence was robust to cardiac motion, and (b) MD was higher than that acquired using standard M2‐nulled LTE, where diffusion‐weighting was applied in three orthogonal directions, which may be attributed to the presence of restricted diffusion and microscopic diffusion anisotropy. Provided adequate signal‐to‐noise ratio, STE could significantly shorten estimation of MD compared with the conventional LTE approach. Importantly, our theoretical analysis and the proposed gradient waveform design may be useful in microstructure imaging beyond diffusion tensor imaging where the effects of motion must be suppressed.     

CTLA4 antagonists in phase I and phase II clinical trials,current status and future perspectives for cancer therapy

Introduction: In cancer, the immune response to tumor antigens is often suppressed by inhibitors and ligands. Checkpoint blockade, considered one of the most promising frontiers for anti-cancer therapy, aims to stimulate the immune anti-cancer response. Agents such as cytotoxic T lymphocyte–associated antigen 4 (CTLA-4) inhibitors offer prolonged survival with manageable side effects.

Areas covered: We summarize the recent clinical successes of CTLA-4 inhibitors and place a strong emphasis on those in early phase clinical trials, often in combination with other immune check-point inhibitors, i.e., programmed cell death protein 1 (PD-1) and BRAF/mitogen-activated protein kinase inhibitors.

Expert opinion: Recent phase I and phase II clinical trials confirm the efficacy of anti-CTLA-4 therapy for treatment of cancers such as renal cell carcinoma. These studies also indicated increased efficacy with combined immune checkpoint blockade with PD-1 or Ras/Raf/mitogen-activated protein kinase/ERK kinase (MEK)/extracellular-signal-regulated kinase (ERK) inhibitors. Researchers must search for new immune targets that may enable more effective and safe immune checkpoint blockade and cancer therapy. This goal may be achieved by next-generation combination therapies to overcome immune checkpoint therapy resistance.     

Contribution of Calcium Channel Subtypes to the Intracellular Calcium Signal in Sensory Neurons: The Effect of Injury

Background: Although the activation-induced intracellular Ca2+ signal is disrupted by sensory neuron injury, the contribution of specific Ca2+ channel subtypes is unknown.

Methods: Transients in dissociated rat dorsal root ganglion neurons were recorded using fura-2 microfluorometry. Neurons from control rats and from neuropathic animals after spinal nerve ligation were activated either by elevated bath K+ or by field stimulation. Transients were compared before and after application of selective blockers of voltage-activated Ca2+ channel subtypes.

Results: Transient amplitude and area were decreased by blockade of the L-type channel, particularly during sustained K+ stimulation. Significant contributions to the Ca2+ transient are attributable to the N-, P/Q-, and R-type channels, especially in small neurons. Results for T-type blockade varied widely between cells. After injury, transients lost sensitivity to N-type and R-type blockers in axotomized small neurons, whereas adjacent small neurons showed decreased responses to blockers of R-type channels. Axotomized large neurons were less sensitive to blockade of N- and P/Q-type channels. After injury, neurons adjacent to axotomy show decreased sensitivity of K+-induced transients to L-type blockade but increased sensitivity during field stimulation.     

Successful regrafting of a transplanted liver

We report on the successful regrafting of a transplanted liver. The donor liver was first grafted into a patient suffering from cryptogenic cirrhosis; the patient died 1 day after the elective transplantation of cerebral bleeding. The well-functioning graft was harvested again and transferred to our institution. After another 12 h of cold ischemia, the liver was reperfused in an urgently registered patient with recurrence of hepatitis B in his first graft. The transplantation was successfully performed and the patient is now doing well, more than 5 months after regrafting with the reused liver. Received: 21 October 1996 Received after revision: 9 January 1997 Accepted: 27 January 1997     

Pituitary responsiveness to GH-releasing hormone, GH-releasing peptide-2 and thyrotrophin-releasing hormone in critical illness

OBJECTIVE Protein hypercatabolism and preservation of fat depots are hallmarks of critical illness, which is associated with blunted pulsatile GH secretion and low circulating IGF-I, TSH, T4 and T3. Repetitive TRH administration is known to reactivate the pituitary-thyroid axis and to evoke paradoxical GH release in critical illness. We further explored the hypothalamic-pituitary function in critical illness by examining the effects of GH-releasing hormone (GHRH) and/or GH-releasing peptide-2 (GHRP-2) and TRH administration. PATIENTS AND DESIGN Critically ill adults (n=40; mean age 55 years) received two i.v. boluses with a 6-hour interval (0900 and 1500 h) within a cross-over design. Patients were randomized to receive consecutively placebo and GHRP-2 (n=10), GHRH and GHRP-2 (n=10), GHRP-2 and GHRH+GHRP-2 (n=10), GHRH+GHRP-2 and GHRH+GHRP-2+TRH (n=10). The GHRH and GHRP-2 doses were 1μg/kg and the TRH dose was 200μg. Blood samples were obtained before and 20, 40, 60 and 120 minutes after each injection. MEASUREMENTS Serum concentrations of GH, T4, T3, rT3, thyroid hormone binding globulin (TBG), IGF-I, insulin and cortisol were measured by RIA; PRL and TSH concentrations were determined by IRMA. RESULTS Critically ill patients presented a striking GH response to GHRP-2 (mean±SEM peak GH 51±9 μg/l in older patients and 102±2μg/l in younger patients; P=0.005 vs placebo). The mean GH response to GHRP-2 was more than fourfold higher than to GHRH (P=0.007). In turn, the mean GH response to GHRH+GHRP-2 was 2.5-fold higher than to GHRP-2 alone (P=0.01), indicating synergism. Adding TRH to the GHRH+GHRP-2 combination slightly blunted this mean response by 18% (P=0.01). GHRP-2 had no effect on serum TSH concentrations whereas both GHRH and GHRH+GHRP-2 evoked an increase in peak TSH levels of 53 and 32% respectively. The addition of TRH further increased this TSH response < ninefold (P=0.005), elicited a 60% rise in serum T3 (P=0.01) and an 18% increase in T4 (P=0.005) levels, without altering rT3 or TBG levels. GHRH and/or GHRP-2 induced a small increase in serum PRL levels. The addition of TRH magnified the PRL response 2.4-fold (P=0.007). GHRP-2 increased basal serum cortisol levels (531±29nmol/l) by 35% (P=0.02); GHRH provoked no additional response, but adding TRH further increased the cortisol response by 20% (P=0.05). CONCLUSIONS The specific character of hypothalamic-pituitary function in critical illness is herewith extended to the responsiveness to GHRH and/or GHRP-2 and TRH. The observation of striking bursts of GH secretion elicited by GHRP-2 and particularly by GHRH+GHRP-2 in patients with low spontaneous GH peaks opens the possibility of therapeutic perspectives for GH secretagogues in critical care medicine.     

Retrograde flush following topical cooling is superior to preserve the non-heart-beating donor lung

Objective: The use of non-heart-beating donors (NHBD) has been propagated as an alternative to overcome the scarcity of pulmonary grafts. Formation of microthrombi after circulatory arrest, however, is a major concern for the development of reperfusion injury. We looked at the effect and the best route of pulmonary flush following topical cooling in NHBD. Methods: Non-heparinized pigs were sacrificed by ventricular fibrillation and divided into three groups (n = 6 per group). After 1 h of in situ warm ischaemia and 2.5 h of topical cooling, lungs in group I were retrieved unflushed (NF). In group II, lungs were explanted following an anterograde flush (AF) through the pulmonary artery with 50 ml/kg Perfadex^® (6 °C). Finally, in group III, lungs were retrieved after an identical but retrograde flush (RF) via the left atrium. Flush effluent was sampled at intervals to measure haemoglobin concentration. Performance of the left lung was assessed during 60 min in our ex vivo reperfusion model. Wet-to-dry weight ratio (W/D) of both lungs was calculated as an index of pulmonary oedema. IL-1ß and TNF- protein levels in bronchial lavage fluid from both lungs were compared between groups. Results: Haemoglobin concentration (g/dl) was higher in the first effluent in RF versus AF (3.4 ± 1.1 vs 0.6 ± 0.1; p < 0.05). Pulmonary vascular resistance (dynes × s × cm⁻⁵) was 975 ± 85 RF versus 1567 ± 98 AF and 1576 ± 88 NF at 60 min of reperfusion (p < 0.001). Oxygenation (mmHg) and compliance (ml/cmH₂O) were higher (491 ± 44 vs 472 ± 61 and 430 ± 33 NS, 22 ± 3 vs 19 ± 3 and 14 ± 1 NS, respectively) and plateau airway pressure (cmH₂O) was lower (11 ± 1 vs 13 ± 1 and 13 ± 1 NS) after RF versus AF and NF, respectively. No differences in cytokine levels or in W/D ratios were observed between groups after reperfusion. Histology demonstrated microthrombi more often present after AF and NF compared to RF. Conclusion: Retrograde flush of the lung following topical cooling in the NHBD results in a better washout of residual blood and microthrombi and subsequent reduced pulmonary vascular resistance upon reperfusion.     

Assessment of virulence of pigeon isolates of Salmonella enterica subsp. enterica serovar typhimurium variant copenhagen for humans

Salmonella enterica serovar Typhimurium variant Copenhagen was isolated from 5 of 152 (3.3%) feral pigeons from the city of Ghent (Belgium) and from 26 pooled fecal samples from 114 pigeon lofts (22.8%). These isolates belonged to phage type (PT) 99. Seven of the pigeon isolates were further compared in vitro to five human variant Copenhagen isolates, 2 isolates of PT 208, 1 isolate each of PT 120 and U302, and a nontypeable isolate. No differences in invasiveness in human intestinal epithelial Caco-2 cells were found. The human strains, however, were able to multiply significantly more inside human THP-1 macrophages than the pigeon strains. After inoculation of mice with a pigeon PT 99 strain, high numbers of Salmonella bacteria were shed with the feces, the internal organs were heavily colonized, and the animals showed severe clinical symptoms resulting in death. In conclusion, the less-pronounced ability of the pigeon variant Copenhagen strains to multiply inside human macrophages than human strains as well as the lack of human PT 99 isolates during 2002, despite the relatively high frequency of this PT in the pigeon population, suggest these strains to be of low virulence to humans. However, the high virulence for mice of the tested strain implies that rodents may act as reservoirs.     

Rescue extracorporeal cardiopulmonary resuscitation in pediatric patients: a nine-year single-center experience in Zagreb,Croatia

AimTo investigate the risk factors and the outcomes of extracorporeal membrane oxygenation (ECMO) in pediatric patients treated at the University Hospital Center Zagreb, the largest center in Croatia providing pediatric ECMO.MethodsThis retrospective study enrolled all the pediatric patients who required E-CPR from 2011 to 2019. Demographic data, cardiac anatomy, ECMO indications, ECMO complications, and neurodevelopmental status at hospital discharge were analyzed.ResultsIn the investigated period, E-CPR was used in 16 children, and the overall survival rate was 37.5%. Six patients were in the neonatal age group, 5 in the infant group, and 5 in the “older” group. There was no significant difference between the sexes. Four patients had an out-of-hospital arrest and 12 had an in-hospital arrest. Twelve out of 16 patients experienced renal failure and needed hemodialysis, with 4 out of 6 patients in the survivor group and 8 out of 10 in the non-survivor group. Survivors and non-survivors did not differ in E-CPR duration time, lactate levels before ECMO, time for lactate normalization, and pH levels before and after the start of ECMO.ConclusionThe similarity of our results to those obtained by other studies indicates that the ECMO program in our hospital should be maintained and improved.

The use of extracorporeal cardiopulmonary resuscitation (E-CPR) is increasing (1). E-CPR is defined as an initiation of extracorporeal membrane oxygenation (ECMO) during active chest compressions. Its main goal is to provide immediate cardiovascular support to patients who do not react to CPR (2) and to lead to survival and a better neurological outcome (3). After administering CPR for more than 30 minutes, survival with conventional CPR measures ranges between 0%-5% (4,5).The most recent systematic review by the International Liaison Committee on Resuscitation from 2015 recommended that E-CPR should be considered for children with underlying cardiac conditions who have an in-hospital cardiac arrest when appropriate protocols, expertise, and equipment are available (6). According to the Extracorporeal Life Support Organization (ELSO) registry from 2017 (7), more than 60 000 people received extracorporeal life support (ECLS), between 2009 and 2015, with an overall survival rate of 61% (7). Pediatric ECMO experience in Slovenia shows that ECMO programs may be incorporated in smaller hospitals in the region (8-10). The ELSO database includes data on all reported pediatric ECMO runs, including those conducted with E-CPR, and in patients with congenital heart surgery and neonates with diaphragmatic hernia or meconium aspiration syndrome, etc. During the 6-year period, 3005 E-CPR runs were reported, with an overall survival to hospital discharge of 43% (7). A survival rate of 31% was reported by Ergűn et al (11) and in E-CPR patients with severe burn injury (12). The longer the CPR duration time, the lower was the survival to discharge rate. Matos et al reported an E-CPR survival-to-discharge rate of 33% after >35 min of chest compressions (13). Other studies reported that the overall survival rate of pediatric E-CPR cases was growing, with better neurological outcomes than among the patients in the CPR group only (14). Pilar et al found that in 73 pediatric cardiac patients requiring cardiopulmonary resuscitation for >30 min (15), the survival to hospital discharge was 43.8%, with 3/4 of the patients having normal neurological function or mild neurological disability (15). Based on ELSO registry, approximately 10% of all ECMO patients meet brain death criteria (7). One of the biggest single-center studies, involving 184 pediatric ECPR patients (16), showed a successful ECMO weaning in 63% of the patients and the overall survival rate to hospital discharge of 43%. In the same study, the risk factors linked to increased mortality were presupport pH<7.1, mechanical complications, and neurological complications (16). The E-CPR use can involve many complications, not necessarily linked to factors preceding cardiac arrest, such as low cardiac output syndrome or irreversible respiratory failure (17). Furthermore, common complications of ECMO treatment are fluid overload and acute kidney injury (18). Many studies showed renal replacement therapy (RRT) to be negatively associated with survival (15,16,18,19).This study assessed the risk factors and the outcomes of ECMO in the largest Croatian center providing pediatric E-CPR experience over nine years and compared the survivor and the non-survivor group.