Can NIRS be a surrogate indicator of elective shunt in carotid endarterectomy? A single-center observational retrospective study says no.

BACKGROUND: Neuromonitoring during carotid endarterectomy (CEA) under general anesthesia is desirable and may be useful for preventing brain ischemia, but the selection of the most appropriate method remains controversial. PURPOSE: To determine the effectiveness of near infrared spectroscopy (NIRS) compared to multimodality intraoperative neuromonitoring (IONM) in indicating elective shunts and predicting postoperative neurological status. METHODS: This is a retrospective observational study including 86 consecutive patients with CEA under general anesthesia. NIRS and multimodality IONM were performed during the procedure. IONM included electroencephalography (EEG), somatosensory evoked potentials (SSEPs) and transcranial motor-evoked potentials (TcMEPs). Sensitivity, specificity, and positive and negative predictive values (PPV and NPV) were calculated for each neuromonitoring modality. RESULTS: NIRS presented a sensitivity and a specificity for detecting brain ischemia of 77.7% and 89.6%, respectively (PPV = 46.6% and NPV = 97.2%). In contrast, a 100% sensitivity and specificity for multimodality IONM was determined (PPV and NPV = 100%). No significant difference (in demographical or clinical data) between "true positive" and "false-positive" patients was identified. Among the methods included in multimodality IONM, EEG showed the best results for predicting postoperative outcome after CEA (PPV and NPV=100%). CONCLUSION: NIRS is inferior to multimodality IONM in detecting brain ischemia and predicting postoperative neurological status during CEA under general anesthesia.

Telomerase and Pluripotency Factors Jointly Regulate Stemness in Pancreatic Cancer Stem Cells.

To assess the role of telomerase activity and telomere length in pancreatic CSCs we used different CSC enrichment methods (CD133, ALDH, sphere formation) in primary patient-derived pancreatic cancer cells. We show that CSCs have higher telomerase activity and longer telomeres than bulk tumor cells. Inhibition of telomerase activity, using genetic knockdown or pharmacological inhibitor (BIBR1532), resulted in CSC marker depletion, abrogation of sphere formation in vitro and reduced tumorigenicity in vivo. Furthermore, we identify a positive feedback loop between stemness factors (NANOG, OCT3/4, SOX2, KLF4) and telomerase, which is essential for the self-renewal of CSCs. Disruption of the balance between telomerase activity and stemness factors eliminates CSCs via induction of DNA damage and apoptosis in primary patient-derived pancreatic cancer samples, opening future perspectives to avoid CSC-driven tumor relapse. In the present study, we demonstrate that telomerase regulation is critical for the "stemness" maintenance in pancreatic CSCs and examine the effects of telomerase inhibition as a potential treatment option of pancreatic cancer. This may significantly promote our understanding of PDAC tumor biology and may result in improved treatment for pancreatic cancer patients.

Anaesthesia and multimodality intraoperative neuromonitoring in carotid endarterectomy. Chronological evolution and effects on intraoperative neurophysiology.

Contingency data was retrospectively collected to evaluate the historical and current ability to provide multimodality intraoperative neurophysiological monitoring during carotid endarterectomy under two conditions: total intravenous anaesthesia (TIVA) and low dose halogenated anaesthesia (SEVO). 229 patients were monitored during carotid endarterectomy procedures under general anaesthesia between 2012 and 2020. 121 Patients were monitored with SEVO at a minimum alveolar concentration less than 0.7 and 108 were monitored using TIVA, according to common anaesthetic practice standards in our hospital across the years. Multimodality IONM was established with electroencephalography, somatosensory evoked potentials and motor evoked potentials. As compared to TIVA, patients monitored with SEVO showed significantly higher motor evoked potential thresholds (313.52 ± 77.74 SEVO and 218.93 V ± 103.2 V TIVA p < 0.05) and lower reproducibility. Electroencephalography and somatosensory evoked potentials showed no significant differences among the groups. When using SEVO, multimodality intraoperative neurophysiological monitoring during carotid endarterectomy could mask or miss a motor isolated change in patients in spite of low dose minimum alveolar concentration and of apparently adequate electroencephalography and somatosensory evoked potentials for monitoring. Given these difficulties, we believe the chronological transfer to TIVA could have improved our ability to establish multimodality intraoperative neurophysiological monitoring during carotid endarterectomy in recent times.

Incidence of surgical site infection in colon surgery according to RENAVE methodology: Prospective study 2017-2019. / Incidencia de infección de localización quirúrgica en cirugía de colon según metodología RENAVE: estudio prospectivo 2017-2019.

BACKGROUND: Navarra Hospital Complex has renovated its healthcare-associated infections surveillance and control methods meeting the requirements of the Spanish National Epidemiologic Surveillance Network. Surgical site infections are one of the most relevant adverse outcomes, being the colon surgery one of the mandatory monitored procedures. This system will ease, not only the yearly estimation of the hospital surgical infection rates, but also its comparison at national and European levels. METHODS: 416 patients underwent surgery between 2017 and 2019. Clinical variables were gathered during the patient hospitalization and up to 30days from surgery, stratifying the cases by their NHSN (National Health Safety Network) surgical infection risk index. A univariant descriptive analysis was performed and outcome indicators were estimated. RESULTS: The cumulative incidence was 10.6%, with 44 cases. The rates were higher among the high-risk subgroups: 25.0% and 42.9%, respectively, for NSHN index categories2 and3. CONCLUSIONS: The incidence was similar to the ones found in other studies carried out in analogous conditions. However, the methodologic variability makes it difficult to compare results.

National Diagnostic Reference Levels for Endovascular Aneurysm Repair and Optimisation Strategies.

OBJECTIVE: The International Commission on Radiological Protection (ICRP) has highlighted the large number of medical specialties using fluoroscopy outside imaging departments without programmes of radiation protection (RP) for patients and staff. Vascular surgery is one of these specialties and endovascular aneurysm repair (EVAR) is one of the most challenging procedures requiring RP guidance and optimisation actions. The recent European Directive on Basic Safety Standards requires the use and regular update of diagnostic reference levels (DRL) for interventional procedures. The objective of the study was to know the doses of patients undergoing EVAR with mobile Xray systems and with hybrid rooms (fixed Xray systems), to obtain national DRLs and suggest optimisation actions. METHODS: The Spanish Chapter of Endovascular Surgery launched a national survey that involved hospitals for 10 autonomous communities representing the 77% of the Spanish population (46.7 million inhabitants). Patient dose values from mobile Xray systems were available from nine hospitals (sample of 165 EVAR procedures) and data from hybrid rooms, from seven hospitals, with dosimetric data from 123 procedures. The initial national DRLs have been obtained, as the third quartile of the median values from the different centres involved in the survey. RESULTS: The proposed national DRLs are 278 Gy cm2 for hybrid rooms and 87 Gy cm2 for mobile Xray systems, and for cumulative air kerma (cumulative AK) at the patient entrance reference point, 1403 mGy for hybrid rooms, and 292 mGy for mobile systems. CONCLUSION: An audit of patient doses for EVAR procedures to identify optimised imaging protocol strategies is needed. It is also appropriate to evaluate the diagnostic information required for EVAR procedures. The increase by a factor of 3.2 (for kerma area product) and 4.8 (for cumulative AK) in the DRLs needs to be justified when the procedures are performed in the hybrid rooms rather than with mobile Xray systems.

Clinical Infections by Herpesviruses in Patients Treated with Valproic Acid: A Nested Case-Control Study in the Spanish Primary Care Database, BIFAP.

The objective of this study is to evaluate the risk of clinical infections by herpesviruses in patients exposed to valproic acid (VPA). We performed a case-control study nested in a primary cohort selected from the Spanish primary care population-based research database BIFAP (Base de datos para la Investigación Farmacoepidemiológica en Atención Primaria) over the period 2001-2015. The events of interest were those diseases caused by any herpesviruses known to infect humans. For each case, up to 10 controls per case matched by age, gender, and calendar date were randomly selected. A conditional logistic regression was used to compute adjusted odds ratios (OR) and their 95% confidence intervals (95% CI). Current use of VPA was associated with a trend towards a reduced risk of clinical infections by herpesviruses as compared with non-users (OR 0.84; CI 95% 0.7-1.0; p = 0.057). Among current users, a trend to a decreased risk with treatment durations longer than 90 days was also observed. The results show a trend to a reduced risk of clinical infection by herpesviruses in patients exposed to VPA. These results are consistent with those in vitro studies showing that, in cultured cells, VPA can inhibit the production of the infectious progeny of herpesviruses. This study also shows the efficient use of electronic healthcare records for clinical exploratory research studies.

Role of Proteolipid Protein in HSV-1 Entry in Oligodendrocytic Cells.

Herpes simplex virus type 1 (HSV-1) has the ability to enter many different hosts and cell types by several strategies. This highly prevalent alphaherpesvirus can enter target cells using different receptors and different pathways: fusion at a neutral pH, low-pH-dependent and low-pH-independent endocytosis. Several cell receptors for viral entry have been described, but several observations suggest that more receptors for HSV-1 might exist. In this work, we propose a novel role for the proteolipid protein (PLP) in HSV-1 entry into the human oligodendrocytic cell line HOG. Cells transfected with PLP-EGFP showed an increase in susceptibility to HSV-1. Furthermore, the infection of HOG and HOG-PLP transfected cells with the R120vGF virus--unable to replicate in ICP4-defficient cells--showed an increase in viral signal in HOG-PLP, suggesting a PLP involvement in viral entry. In addition, a mouse monoclonal antibody against PLP drastically inhibited HSV-1 entry into HOG cells. PLP and virions colocalized in confocal immunofluorescence images, and in electron microscopy images, which suggest that PLP acts at the site of entry into HOG cells. Taken together these results suggest that PLP may be involved in HSV-1 entry in human oligodendrocytic cells.

The effect of cellular differentiation on HSV-1 infection of oligodendrocytic cells.

Herpes simplex type 1 (HSV-1) is a neurotropic virus that infects many types of cells. Previous studies have demonstrated that oligodendrocytic cells are highly susceptible to HSV-1 infection. Here we analysed HSV-1 infection of a human oligodendrocytic cell line, HOG, and oligodendrocyte precursor cells (OPCs) cultured under growth or differentiation conditions. In addition to cell susceptibility, the role of the major cell receptors for viral entry was assessed. Our results revealed that OPCs and HOG cells cultured under differentiation conditions became more susceptible to HSV-1. On the other hand, viral infection induced morphological changes corresponding to differentiated cells, suggesting that HSV-1 might be inducing cell differentiation. We also observed colocalization of HVEM and nectin-1 with viral particles, suggesting that these two major HSV-1 receptors are functional in HOG cells. Finally, electron microscopy assays indicated that HSV-1 may be also entering OLs by macropinocytosis depending on their differentiation stage. In addition, vesicles containing intracellular enveloped virions observed in differentiated cells point to an endocytic mechanism of virus entry. All these data are indicative of diverse entry pathways dependent on the maturation stage of OLs.

Evidence of Muller's ratchet in herpes simplex virus type 1.

Population bottlenecks can have major effects in the evolution of RNA viruses, but their possible influence in the evolution of DNA viruses is largely unknown. Genetic and biological variation of herpes simplex virus type 1 (HSV-1) has been studied by subjecting 23 biological clones of the virus to 10 plaque-to-plaque transfers. In contrast to large population passages, plaque transfers led to a decrease in replicative capacity of HSV-1. Two out of a total of 23 clones did not survive to the last transfer in 143 TK(-) cells. DNA from three genomic regions (DNA polymerase, glycoprotein gD and thymidine kinase) from the initial and passaged clones was sequenced. Nucleotide substitutions were detected in the TK and gD genes, but not in the DNA polymerase gene. Assuming a uniform distribution of mutations along the genome, the average rate of fixation of mutations was about five mutations per viral genome and plaque transfer. This value is comparable to the range of values calculated for RNA viruses. Four plaque-transferred populations lost neurovirulence for mice, as compared with the corresponding initial clones. LD(50) values obtained with the populations subjected to serial bottlenecks were 4- to 67-fold higher than for their parental clones. These results equate HSV-1 with RNA viruses regarding fitness decrease as a result of plaque-to-plaque transfers, and show that population bottlenecks can modify the pathogenic potential of HSV-1. Implications for the evolution of complex DNA viruses are discussed.

Role of the small GTPase Rab27a during herpes simplex virus infection of oligodendrocytic cells.

BACKGROUND: The morphogenesis of herpes simplex virus type 1 (HSV-1) comprises several events, of which some are not completely understood. It has been shown that HSV-1 glycoproteins accumulate in the trans-Golgi network (TGN) and in TGN-derived vesicles. It is also accepted that HSV-1 acquires its final morphology through a secondary envelopment by budding into TGN-derived vesicles coated with viral glycoproteins and tegument proteins. Nevertheless, several aspects of this process remain elusive. The small GTPase Rab27a has been implicated in regulated exocytosis, and it seems to play a key role in certain membrane trafficking events. Rab27a also seems to be required for human cytomegalovirus assembly. However, despite the involvement of various Rab GTPases in HSV-1 envelopment, there is, to date, no data reported on the role of Rab27a in HSV-1 infection. RESULTS: Herein, we show that Rab27a colocalized with GHSV-UL46, a tegument-tagged green fluorescent protein-HSV-1, in the TGN. In fact, this small GTPase colocalized with viral glycoproteins gH and gD in that compartment. Functional analysis through Rab27a depletion showed a significant decrease in the number of infected cells and viral production in Rab27a-silenced cells. CONCLUSIONS: Altogether, our results indicate that Rab27a plays an important role in HSV-1 infection of oligodendrocytic cells.

Glycoprotein G from pseudorabies virus binds to chemokines with high affinity and inhibits their function.

Pseudorabies virus (PRV), also known as suid herpesvirus, is the aetiological agent of Aujeszky's disease in swine. In other animals, except higher-order primates, PRV infection is often fatal. The mechanisms of PRV pathogenesis and immune modulation are largely unknown. PRV codes for 11 glycoproteins. Among them, glycoprotein G (gG) is the most abundant PRV protein found in the supernatant of PRV-infected cell cultures. PRV-gG has low amino acid sequence similarity with gG from other animal alphaherpesviruses and its function is unknown. gG from other animal alphaherpesviruses, with the exception of at least equine herpesvirus 4, binds to chemokines. We show here that PRV-gG binds to the human chemokine CL1 and several CC and CXC human chemokines with high affinity. Chemokine-binding activity can be detected in the supernatants of PRV-infected cell cultures, and insertional inactivation of the gene encoding gG from the PRV genome results in loss of chemokine-binding activity. Binding of PRV-gG to chemokines inhibits chemokine-mediated cell migration, suggesting a role for PRV-gG in immune evasion.

High susceptibility of a human oligodendroglial cell line to herpes simplex type 1 infection.

More than 20 infectious agents, ranging from retroviruses to mycobacteria, have been associated with multiple sclerosis onset or relapses in which oligodendrocytes, the myelin-forming cells of the central nervous system, are the initial target of the pathogenic status. In this work, the nature of the susceptibility of the human precursor oligodendroglial KG-1C cell line to herpes simplex virus type 1 (HSV-1) was investigated. Infection of KG-1C cells was characterized by a high level of virus production and a notable progression of the cytopathic effect. After infection, there was a significant shut-off of host mRNA translation, which was correlated with evident synthesis of viral proteins. An examination by electron microscopy of the infected cells revealed the presence of large clusters of mitochondria located in the proximity of intracellular HSV-1 particle groups. In addition, transmission electron microscopy and nuclear fluorescence analysis showed neither signs of chromatin condensation nor of apoptotic bodies. Furthermore, procaspase-3 remained uncleaved, suggesting that apoptosis does not take place, at least in this system. Finally, expression and localization of MAL2, a subpopulation of detergent-insoluble lipid raft protein, was studied. Detection of MAL2 significantly increased after infection and it was colocalized with HSV-1 proteins. From these findings the authors conclude that human oligodendrocyte-like cells are highly susceptible to HSV-1 infection. The implications of this for central nervous system viral infection are discussed.

Eimeria tenella sporozoites and merozoites differentially express glycosylphosphatidylinositol-anchored variant surface proteins.

Little is known about glycosylphosphatidylinositol (GPI)-linked surface proteins in the coccidian parasite Eimeria tenella. Examination of 28,550 EST sequences from the sporozoite and second merozoite developmental stages of the parasite led to the identification of 37 potential GPI-linked variant surface proteins, termed EtSAGs. Analysis of the complete nucleotide sequences of 23 EtSAG genes separated them into two multi-gene families. All the predicted EtSAG proteins (which vary in length from 228 to 271 residues) have an N-terminal hydrophobic signal peptide, a C-terminal hydrophobic GPI signal-anchor peptide and an extracellular domain organised around six cysteine residues, the positions of which are conserved within each family. Using specific antibodies against a small number of recombinant-expressed EtSAGs, the surface localisation and GPI-anchorage of members of both families was confirmed experimentally. Expression of EtSAGs is differentially regulated between the oocyst/sporozoite and second generation merozoite stages, with only one expressed specifically in the sporozoite, a small number expressed in both stages and the majority expressed specifically in the second generation merozoite. Preliminary data support a model in which multiple variant surface antigens are co-expressed on individual parasites, rather than a model of antigenic switching. The biological role(s) of EtSAGs and the effect(s) that expression of a complex repertoire of variant surface antigens by the second generation merozoite has on host adapted immunity are unknown.

Characterisation of penciclovir resistant acyclovir sensitive herpes simplex virus type 2 isolated from an AIDS patient.

A heterogeneous herpes simplex virus type 2 (HSV-2) population was characterised from an AIDS patient with relapsing genital ulcer. The isolate had an unusual antiviral spectrum, showing resistance to penciclovir and susceptibility to acyclovir. Two viral populations were plaque purified, one resistant and the other susceptible to both antiviral drugs. The resistant clone was deficient in thymidine kinase (TK) activity and a nucleotide substitution, thymine for cytosine, at position 153 was identified in its TK gene. This mutation resulted in an amino acid change, arginine to tryptophan, in the ATP binding site. In the deficient mutant, a loss of virulence was observed in mice.

Upper arm polytetrafluoroethylene grafts for dialysis access. Analysis of two different graft sizes: 6 mm and 6-8 mm.

The purpose of this retrospective study was to analyze 2 sizes of expanded polytetrafluoroethylene (PTFE) upper arm grafts for dialysis: 8 millimeters, tapered to 6 mm at the arterial side, and 6 millimeters. All upper arm PTFE grafts (Gore-Tex(R)) were performed between January 1981 and April 1997. Patient characteristics, complication rate, and patency rates were compared for both kind of grafts. Five hundred and seven PTFE grafts were analyzed (183 6-mm grafts and 324 6- to 8-mm grafts). Early failure was found in 5 grafts (2.7%) in 6-mm grafts, and in 5 grafts (1.5%) in 8-mm grafts (not significant). Steal syndrome was found in 1 patient (0.5%) of the 6-mm group, and in 11 (3.4%) of the 8-mm grafts (p=0.085). The rate of late complications requiring surgical repair was 0.56 episode per graft-year in the 6-mm grafts group, and 0.33 in the 8-mm grafts (p<0.001). Primary patency rates of 6-mm grafts were 72%, 33%, and 19% at 1, 3, and 5 years; and secondary patency rates were 86%, 68%, 56%, and 44% at 1, 3, 5, and 6 years, respectively. In the 8-mm grafts group, primary patency rates were 77%, 52%, and 39% at 1, 3, and 5 years; and secondary patency rates were 92%, 84%, 73%, and 66% at 1, 3, 5, and 6 years, respectively. Comparison of patency rates of 6-mm and 8-mm grafts were statistically significant (p<0.001) for both primary and secondary curves. However, secondary survival curves were similar for both kind of grafts in a subpopulation of diabetic patients. The authors conclude that the 8-mm graft, tapered to 6 mm at the arterial side, is a dialysis graft with fewer complications and a better patency rate than grafts of 6 mm placed in the same anatomical position, at least in a population of nondiabetic patients. Steal syndrome was observed in some cases of diabetic and older patients with a large-bore graft. Thus, this kind of prosthesis should be avoided in this population. On the other hand, this is not a prospective, randomized study made with any intention for comparison. Therefore, the aforementioned conclusions must be cautiously considered.

Development of new expression vector based on Pseudorabies virus amplicons: application to human insulin expression.

Pseudorabies virus (PrV), a herpesvirus from the Alphaherpesvirinae subfamily, is suitable for amplicon vector replication and packaging into virions, with helper virus for trans replication and cleavage-packaging functions. PrV amplicon vectors were developed in a bacterial plasmid construction using PrV ori(s) and pac signals as the required cis elements. Human insulin cDNA was then cloned in the amplicon vector for human proinsulin expression. In the same construction, green fluorescent protein was used as a marker. PrV amplicons may have several advantages over herpes simplex virus type 1 (HSV1) amplicons in human gene therapy because it can infect human cells in vitro and in vivo, it is not pathogenic for primates and there is no pre-existing immunity and risk of recombination with latent PrV as occurs with HSV1.

Characterization of regions in the GP5 protein of porcine reproductive and respiratory syndrome virus required to induce apoptotic cell death.

Expression of the GP5 protein of porcine reproductive and respiratory syndrome virus in mammalian cells using a recombinant vaccinia virus has been shown to induce strong cytotoxicity due to apoptotic death. We have now developed a transient expression system that allows the observation and quantitation of the cell death due to GP5 synthesis, taking advantage of the reduction that this protein induces in the expression of two different co-transfected reporter genes. In this way, we are able to study the regions in GP5 implicated in apoptosis induction. The first 119 aminoacids constitute a region capable of fully inducing apoptosis, aminoacids 90-119 having a fundamental role. On the contrary, the C-terminal region is unable by itself of cell death induction and, moreover, is dispensable for this phenotype. We have also observed that induction of apoptosis is independent of cleavage of the N-terminal putative signal sequence in GP5 or N-glycosylation of this protein.