Estimation of the burden of flu-association influenza-like illness visits on total clinic visits through the sentinel influenza monitoring system in Senegal during the 2013-2015 influenza seasons.

Knowing the burden of influenza is helpful for policy decisions. Here we estimated the contribution of influenza-like illness (ILI) visits associated with laboratory-confirmed influenza among all clinic visits in a Senegal sentinel network. ILI data from ten sentinel sites were collected from January 2013 to December 2015. ILI was defined as an axillary measured fever of more than 37.5 °C with a cough or a sore throat. Collected nasopharyngeal swabs were tested for influenza viruses by rRT-PCR. Influenza-associated ILI was defined as ILI with laboratory-confirmed influenza. For the influenza disease burden estimation, we used all-case outpatient visits during the study period who sought care at selected sites. Of 4030 ILI outpatients tested, 1022 were influenza positive. The estimated proportional contribution of influenza-associated ILI was, per 100 outpatients, 1.2 (95% CI 1.1-1.3), 0.32 (95% CI 0.28-0.35), 1.11 (95% CI 1.05-1.16) during 2013, 2014, 2015, respectively. The age-specific outpatient visits proportions of influenza-associated ILI were higher among children under 5 years (0.68%, 95% CI: 0.62-0.70). The predominant virus during years 2013 and 2015 was influenza B while A/H3N2 subtype was predominant during 2014. Influenza viruses cause a substantial burden of outpatient visits particularly among children under 5 of age in Senegal and highlight the need of vaccination in risk groups.

Ninety Seconds Could be the Optimal Duration for Ventricular Radiofrequency Ablation - Results From a Myocardial Phantom Model.

BACKGROUND: Shallow lesions could be the predominant factor affecting the efficacy of ventricular radiofrequency (RF) ablations. The objective of this study was to assess lesion dimensions and overheating in extended RF ablations up to 180seconds and compare with that of conventional 30seconds ablations. METHODS: The Navistar Thermocool irrigated catheter (Biosense Webster, CA, USA) was used in a previously validated myocardial phantom. Ablations were performed with 20W, 30W, 40W and 50W powers for 180seconds. The volume of lesion and overheating were measured at 530C and 800C isotherms respectively. RESULTS: A total of 110 RF lesions were analysed. The lesion depth increment when ablation was extended from the conventional 30seconds to 90seconds were 31.2±0.2, 33.6±0.6, 36.3±1.8% of that at 30seconds, respectively for powers 30W, 40W and 50W. During 30W ablations, at 90seconds the lesion width and depth were 95.4±1.2%, 91.8±1.6% respectively of the final dimensions at 180seconds. Similar proportions were observed for 40W and 50W. During 40W ablations, the volume of overheating was 113±6% and 184±11% higher at 90seconds and 180seconds respectively compared to that at 30seconds and was 142±9% and 194±9% for 50W ablations. CONCLUSION: Extending RF ablations up to 90seconds significantly increased the lesion depth (30-40%), however, overheating was present at 40W and 50W powers. Ablations beyond 90seconds provided little incremental value.

Circuit Impedance Could Be a Crucial Factor Influencing Radiofrequency Ablation Efficacy and Safety: A Myocardial Phantom Study of the Problem and its Correction.

BACKGROUND: Circuit impedance could affect the safety and efficacy of radiofrequency (RF) ablation. AIM: To perform irrigated RF ablations with graded impedance to compare (1) lesion dimensions and overheated dimensions in fixed power ablations (2) and in power corrected ablations. METHODS: Ablations were performed with irrigated Navistar Thermocool catheter and Stockert EP shuttle generator at settings of 40 W power for 60 seconds, in a previously validated myocardial phantom. The impedance of the circuit was set at 60 Ω, 80 Ω, 100 Ω, 120 Ω, 140 Ω, and 160 Ω. The lesion and overheated dimensions were measured at 53 °C and 80 °C isotherms, respectively. In the second set of ablations, power was corrected according to circuit impedance. RESULTS: In total, 70 ablations were performed. The lesion volume was 72.0 ± 4.8% and 44.7 ± 4.6% higher at 80 Ω and 100 Ω, respectively, compared to that at 120 Ω and it was 15.4 ± 1.2%, 28.1 ± 2.0%, and 38.0 ± 1.8% lower at 140 Ω, 160 Ω, and 180 Ω, respectively. The overheated volume was four times larger when impedance was reduced to 80 Ω from 100 Ω. It was absent at 120 Ω and above. In the power corrected ablations, the lesion volumes were similar to that of 40 W/120 Ω ablations and there was no evidence of overheating. CONCLUSION: The lesion and overheated dimensions were significantly larger with lower circuit impedance during irrigated RF ablation and the lesion size was smaller in high impedance ablations. Power delivery adjusted to impedance using a simple equation improved the consistency of lesion formation and prevented overheating.

In vivo evaluation of virtual electrode mapping and ablation utilizing a direct endocardial visualization ablation catheter.

BACKGROUND: Radiofrequency (RF) ablation utilizing direct endocardial visualization (DEV) requires a "virtual electrode" to deliver RF energy while preserving visualization. This study aimed to: (1) examine the virtual electrode RF ablation efficacy; (2) determine the optimal power and duration settings; and (3) evaluate the utility of virtual electrode unipolar electrograms. METHODS AND RESULTS: The DEV catheter lesions were compared to lesions formed using a 3.5 mm open irrigated tip catheter within the right atria of 12 sheep. Generator power settings for DEV were titrated from 12W, 14W and 16W for 20, 30 and 40 seconds duration with 25 mL/min saline irrigation. Standard irrigated tip catheter settings of 30W, 50°C for 30 seconds and 30 mL/min were used. The DEV lesions were significantly greater in surface area and both major and minor axes compared to irrigated tip lesions (surface area 19.43 ± 9.09 vs 10.88 ± 4.72 mm, P<0.01) with no difference in transmurality (93/94 vs 46/47) or depth (1.86 ± 0.75 vs 1.85 ± 0.57 mm). Absolute electrogram amplitude reduction was greater for DEV lesions (1.89 ± 1.31 vs 1.49 ± 0.78 mV, P = 0.04), but no difference in percentage reduction. Pre-ablation pacing thresholds were not different between DEV (0.79 ± 0.36 mA) and irrigated tip (0.73 ± 0.25 mA) lesions. There were no complications noted during ablation with either catheter. CONCLUSIONS: Virtual electrode ablation consistently created wider lesions at lower power compared to irrigated tip ablation. Virtual electrode electrograms showed a comparable pacing and sensing efficacy in detecting local myocardial electrophysiological changes.

Toward cell-targeting gene therapy vectors: selection of cell-binding peptides from random peptide-presenting phage libraries.

Ideal gene therapy vectors would be delivered intravenously to transfect only specific cells. Existing vectors only transfect cells in vivo in a manner determined by blood flow and the site of introduction. As a general and systematic approach for generating cell-targeting ligands for gene therapy vectors, we have used peptide-presenting phage libraries to select peptides that bind and enter several different cell types. Because of their small size, cell-binding peptides such as these could be incorporated into biological or physical gene therapy vectors. In addition, peptide-presenting phage themselves may also be candidates for gene therapy vectors.

Semaphorin III can repulse and inhibit adult sensory afferents in vivo.

During development, semaphorins (collapsin, fasciclin) mediate repulsive and inhibitory guidance of neurons. Semaphorin III, a secretable member of this family, is expressed by the ventral spinal cord at the time corresponding to projection of sensory afferents from the dorsal root ganglion (DRG) into the spinal cord. The inhibitory effect of E14 ventral cord is active only on nerve growth factor (NGF)-responsive sensory afferents (small-diameter A-delta and C fibers subserving sensations of temperature and pain). Similarly, COS cells secreting recombinant semaphorin III are able to selectively repel DRG afferents whose growth is stimulated by NGF and not NT-3. However, it is not known whether these molecules can exert a functional role in the fully developed adult peripheral nervous system. In this study, we demonstrated that gene gun transfection and production of semaphorin III in corneal epithelial cells in adult rabbits in vivo can cause repulsion of established A-delta and C fiber trigeminal sensory afferents. In addition, it is shown that, following epithelial wounding and denervation, semaphorin III is able to inhibit collateral nerve sprouts from innervating the reepithelialized tissue. These findings are significant in that they provide direct evidence that small-diameter adult sensory neurons retain the ability to respond to semaphorin III. In addition, the corneal gene gun technique may be generally used to study the in vivo effects of neural growth modulators by quantifying the amount of sensory nerve growth.

CD4+ T-lymphopenia in HIV negative tuberculous patients at King Khalid University Hospital in Riyadh, Saudi Arabia.

Tuberculosis (Tb) is a chronic infectious disease in which the cellular immunity (specifically CD4+ and CD8 lymphocytes) provides the most important defense in controlling infection. CD4 lymphopenia is a well-defined risk factor for the development of active tuberculosis in patients infected with Human Immunodeficiency Virus. In HIV - negative patients, CD4 and CD8 cell count suppression has been associated with Tb infection. Our study was designed to determine the baseline and post-treatment values of CD4 and CD8 in HIV negative patients diagnosed with active Tb in Saudi Arabian patients. We recruited twenty-eight, non-HIV patients with tuberculosis for the study group comprising 16 males and 12 females with either disseminated or localized active Tb infection. Two control groups were selected one of twenty-one matched healthy controls and the second of forty-two subjects from pool of controls of an ongoing study in same population for normal CD4 and CD8 counts. The baseline pre-treatment CD4 and CD8 counts in the study group were significantly lower than either control group. Specifically the mean ± SD of CD4 counts were 556.79 ± 298.81 in the study group vs 1,132.38 ± 259.90 in control group 1 and 1,424.38 ± 870.98 in control group 2 (p 0.000). Likewise the CD8 counts in the study group were 1,136.00 ± 512.06 vs. 1,461.90 ± 367.02 in control group 1 and 1,495.90 ± 565.32 in control group 2 (p 0.000) respectively. After treatment of tuberculosis, the study patients experienced a significant increase in their mean ± SD CD4 and CD8 cell counts, from 556.79 ± 297.81 to 954.29 ± 210.90 for CD4 cells (p 0.005) and 1136.00 ± 512.06 to 1,316.54 ± 286.17 for CD8 cells (p 0.002). Analysis of study patients with disseminated disease found significantly lower CD4 cells (but not lower CD8 cells) compared to study patients with localized disease, both at baseline and after treatment. The mean ± SD baseline CD4 cells were 247.60 ± 187.80 with disseminated vs 728.56 ± 186.32 for localized disease (p = 0.000) which rose to 842.30 ± 93.55 vs 1016.50 ± 233.51 (p = 0.033) respectively. We conclude that tuberculosis may be associated with CD4 and CD8 lymphopenia even in patients without human immunodeficiency virus infection, there was the tendency of recovery towards normality especially of the CD4 and CD8 counts after treatment, and that disseminated disease is associated specifically with profound CD4 lymphopenia.

Adenovirus serotype 5 hexon is critical for virus infection of hepatocytes in vivo.

Human species C adenovirus serotype 5 (Ad5) is the most common viral vector used in clinical studies worldwide. Ad5 vectors infect liver cells in vivo with high efficiency via a poorly defined mechanism, which involves virus binding to vitamin K-dependent blood coagulation factors. Here, we report that the major Ad5 capsid protein, hexon, binds human coagulation factor X (FX) with an affinity of 229 pM. This affinity is 40-fold stronger than the reported affinity of Ad5 fiber for the cellular receptor coxsackievirus and adenovirus receptor, CAR. Cryoelectron microscopy and single-particle image reconstruction revealed that the FX attachment site is localized to the central depression at the top of the hexon trimer. Hexon-mutated virus bearing a large insertion in hexon showed markedly reduced FX binding in vitro and failed to deliver a transgene to hepatocytes in vivo. This study describes the mechanism of FX binding to Ad5 and demonstrates the critical role of hexon for virus infection of hepatocytes in vivo.

The spiracular organ of sharks and skates: anatomical evidence indicating a mechanoreceptive role.

The elasmobranch spiracular organ is a specialized receptor associated with the first visceral pouch. The structure of the sensory epithelium of the spiracular organ and the pattern of central termination of the afferent neurons that innervate it show that the spiracular organ is a mechanoreceptor closely related to the lateral line system of sense organs. Its position and orientation within the spiracular cleft suggest that it plays a role in proprioception or equilibrium-audition.

Quantitative spectral assessment of intracardiac electrogram characteristics associated with post infarct fibrosis and ventricular tachycardia.

BACKGROUND: Post-myocardial infarction (MI) remodeling contributes to increased electrophysiological and structural heterogeneity and arrhythmogenesis. Utilising the post-infarct ovine model our aim was to determine unipolar electrogram frequency characteristics consequent to this remodeling and the development of Ventricular Tachycardia (VT). METHODS AND RESULTS: Mapping studies were performed on 14 sheep at >1 month post-MI induction. Sheep were divided into VT inducible (n = 7) and non-inducible (n = 7) groups. Multielectrode needles (n = 20) were deployed within and surrounding ventricular scar for electrophysiological assessment of electrogram amplitude and width. Spectral analysis of electrograms was undertaken using wavelet and fast fourier transformations (WFFT) to calculate root mean square (RMS) power intervals spanning 0-300Hz in 20Hz intervals. Quantitative assessment between electrophysiological and histological parameters including collagen density, and structural organization of the myocardium was performed. Increasing myocardial scar density resulted in attenuation of electrogram amplitude and RMS values. (all p<0.01). Between groups there were no differences in electrogram amplitude (p = 0.37), however WFFT analysis revealed significantly higher RMS values in the VT group (p<0.05) in association with high frequency fractional components of the electrogram. As scar density increased, greater between-group differences in RMS were observed spanning this high frequency (200-280Hz) spectrum and which were proportionally dependent on the degree of structural disorganisation of the myocardium (p<0.001) and number of extrastimuli required to induce VT (p<0.05). CONCLUSION: High frequency unipolar electrogram spectral characteristics were quantitatively co-influenced by the presence of fibrosis and degree of myocardial structural dissorganisation and were associated with the propensity for development of VT.

Analysis of events associated with cell cycle arrest at G2 phase and cell death induced by cisplatin.

DNA is the accepted target for cisplatin, but recent evidence has shed doubt on DNA synthesis as the critical process. L1210/0 cells incubated for 2 hours with cisplatin progress to the G2 phase of the cell cycle and are arrested there for several days. They then either progress in the cell cycle or die. In cells that eventually die, total transcription, polyadenylated [poly(A)+] RNA synthesis, and protein synthesis were markedly inhibited only after 48 hours. Nicotinamide adenine dinucleotide (NAD) and adenosine triphosphate (ATP) levels decreased after 3 days. Cell membrane integrity was lost after 4 days. These results demonstrate that cells can be lethally damaged, yet continue to undergo apparently normal metabolic activities for several days. In a previous study, DNA double-strand breaks were detected after 1 day. We now show that by 2 days, breaks are visible as fragmentation in the nucleosome spacer regions of chromatin. This type of damage is consistent with cell death occurring by the process of apoptosis. Cell shrinkage and morphology were also consistent with this type of cell death. The slow cell death reported here appears to occur at the G2/M transition and may involve events that normally occur at this stage of the cell cycle. These results demonstrate the importance of DNA degradation as an early and possibly essential step in cell death.

Etoposide-induced apoptosis in human HL-60 cells is associated with intracellular acidification.

Apoptosis is a pathway of cell death characterized by internucleosomal digestion of genomic DNA. Such DNA digestion can be induced by both physiological stimuli and cytotoxic treatment with many anticancer agents. This digestion has generally been considered to be mediated by a Ca2+/Mg(2+)-dependent endonuclease that is activated by increases in intracellular Ca2+. However, we suggest that an alternate endonuclease, DNase II, may be a more likely candidate. In these studies, apoptosis was induced in human HL-60 cells by a 30-min incubation with the topoisomerase II inhibitor etoposide. DNA digestion characteristic of apoptosis began within 3 h of removal of etoposide. Morphological indication of apoptosis was observed concurrently. Only about 20% of the cells underwent apoptosis at this time; these appeared to be cells in S phase at the time of etoposide treatment. The remainder of the cells progressed to the G2 phase and arrested there for at least 48 h. Intracellular Ca2+ and pH were measured in individual cells by flow cytometry. No changes in intracellular Ca2+ were observed, but an acidification of up to 1 pH unit occurred in about 15% of the cells and correlated with the time course of appearance of DNA digestion. Cells were sorted on the basis of intracellular pH and only the acidic cells showed the morphology and DNA digestion characteristic of apoptosis. These results demonstrate the involvement of DNase II in apoptotic DNA digestion and suggest mechanisms of pH homeostasis as regulators of apoptosis.

In vivo evidence of mitochondrial dysfunction and altered redox homeostasis in a genetic mouse model of propionic acidemia: Implications for the pathophysiology of this disorder.

Accumulation of toxic metabolites has been described to inhibit mitochondrial enzymes, thereby inducing oxidative stress in propionic acidemia (PA), an autosomal recessive metabolic disorder caused by the deficiency of mitochondrial propionyl-CoA carboxylase. PA patients exhibit neurological deficits and multiorgan complications including cardiomyopathy. To investigate the role of mitochondrial dysfunction in the development of these alterations we have used a hypomorphic mouse model of PA that mimics the biochemical and clinical hallmarks of the disease. We have studied the tissue-specific bioenergetic signature by Reverse Phase Protein Microarrays and analysed OXPHOS complex activities, mtDNA copy number, oxidative damage, superoxide anion and hydrogen peroxide levels. The results show decreased levels and/or activity of several OXPHOS complexes in different tissues of PA mice. An increase in mitochondrial mass and OXPHOS complexes was observed in brain, possibly reflecting a compensatory mechanism including metabolic reprogramming. mtDNA depletion was present in most tissues analysed. Antioxidant enzymes were also found altered. Lipid peroxidation was present along with an increase in hydrogen peroxide and superoxide anion production. These data support the hypothesis that oxidative damage may contribute to the pathophysiology of PA, opening new avenues in the identification of therapeutic targets and paving the way for in vivo evaluation of compounds targeting mitochondrial biogenesis or reactive oxygen species production.

A foodborne outbreak of Cyclospora cayetanensis at a wedding: clinical features and risk factors for illness.

BACKGROUND: Cyclospora cayetanensis, a coccidian parasite, has increasingly been recognized as a cause of gastrointestinal tract illness. We describe an outbreak of Cyclospora infection following a wedding reception. OBJECTIVES: To investigate and characterize risk factors associated with the outbreak of Cyclospora and to describe the observed clinical course and spectrum of illness. METHODS: Retrospective cohort study involving 94 of the 101 guests who attended a wedding reception at a restaurant in Boston, Mass. RESULTS: Fifty-seven respondents met the case definition of infection; 12 of these had laboratory-confirmed Cyclospora. The epidemic curve was consistent with a point source outbreak with a median incubation period of 7 days. Commonly reported symptoms included diarrhea (100%), weight loss (93%), fatigue (91%), and anorexia (90%). The illness had a characteristic waxing and waning course, with 51 persons (89%) reporting recurring symptoms and 35 (61%) reporting illness lasting more than 3 weeks. By univariate analysis, infection was significantly associated (P<.05) with consumption of wine and a dessert containing raspberries, strawberries, blackberries, and blueberries. Only the dessert remained significant by stratified analysis with an adjusted relative risk of 2.1 (95% confidence interval, 1.4-3.2). CONCLUSIONS: Findings from this study support a point source outbreak of the newly identified pathogen C cayetanensis, with berries as the vehicle of transmission. It suggests that Cyclospora may cause severe diarrhea associated with profound anorexia and weight loss, and should be considered in the evaluation of prolonged gastrointestinal tract illness.

Nosocomial infection and fatality in medical and surgical intensive care unit patients.

We prospectively studied 526 patients admitted to the medical intensive care unit (MICU) and 799 patients admitted to the surgical intensive care unit (SICU) at a municipal hospital over a 20-month period. Rates of nosocomial infection were higher in the SICU patients (31% vs 24%). The SICU patients had more urinary tract infections, bacteremias, and wound infections, and the MICU patients were older, had higher acute physiology scores on admission and were more often admitted with shock or coma. The SICU patients were more likely to have received prior antibiotic therapy and had significantly higher numbers of endotracheal tubes, arterial lines, central venous lines, and indwelling bladder catheters. Of the 23 variables univariately associated with nosocomial infection, only five remained significant after entry into step-wise regression models. The MICU patients had a higher fatality rate in the MICU than did the SICU patients (18% vs 10%), but the relative risk of a death following nosocomial infection was 3.5 for both groups. Thirty variables were significantly associated with hospital fatality; nine remained significant after analysis by stepwise logistic regression.

Perfusion redistribution after a pulmonary-embolism-like event with contrast enhanced EIT.

Recent studies showed that regional pulmonary perfusion can be reliably estimated using electrical impedance tomography (EIT) with the aid of hypertonic saline based contrast enhancement. Building on these successful studies, we studied contrast EIT for pulmonary perfusion defect caused by an artificially induced pulmonary embolism (PE) in a large ovine model (N = 8, 78 ± 7.8 kg). Furthermore, the efficacy of a less invasive contrast bolus of 0.77 ml kg(-1) of NaCl 3% was compared with a more concentrated bolus of 0.13 ml kg(-1) of NaCl 20%. Prior to the injection of each contrast bolus injection, ventilation was turned off to provide a total of 40 to 45 s of apnoea. Each bolus of impedance contrast was injected through a catheter into the right atrium. Pulmonary embolisation was performed by balloon occlusion of part of the right branch of the pulmonary trunk. Four parameters representing the kinetics of the contrast dilution in the lung were evaluated for statistical differences between baseline and PE, including peak value, maximum uptake, maximum washout and area under the curve of the averaged contrast dilution curve in each lung. Furthermore, the right lung to left lung (R2L) ratio of each the aforementioned parameters were assessed. While all of the R2L ratios yielded significantly different means between baseline and PE, it can be concluded that the R2L ratios of area under the curve and peak value of the averaged contrast dilution curve are the most promising and reliable in assessing PE. It was also found that the efficacy of the two types of impedance contrasts were not significantly different in distinguishing PE from baseline in our model.

Magnetic guidance versus manual control: comparison of radiofrequency lesion dimensions and evaluation of the effect of heart wall motion in a myocardial phantom.

BACKGROUND: Magnetic navigation system (MNS) ablation was suspected to be less effective and unstable in highly mobile cardiac regions compared to radiofrequency (RF) ablations with manual control (MC). The aim of the study was to compare the (1) lesion size and (2) stability of MNS versus MC during irrigated RF ablation with and without simulated mechanical heart wall motion. METHODS: In a previously validated myocardial phantom, the performance of Navistar RMT Thermocool catheter (Biosense Webster, CA, USA) guided with MNS was compared to manually controlled Navistar irrigated Thermocool catheter (Biosense Webster, CA, USA). The lesion dimensions were compared with the catheter in inferior and superior orientation, with and without 6-mm simulated wall motion. All ablations were performed with 40 W power and 30 ml/ min irrigation for 60 s. RESULTS: A total of 60 ablations were performed. The mean lesion volumes with MNS and MC were 57.5 ± 7.1 and 58.1 ± 7.1 mm(3), respectively, in the inferior catheter orientation (n = 23, p = 0.6), 62.8 ± 9.9 and 64.6 ± 7.6 mm(3), respectively, in the superior catheter orientation (n = 16, p = 0.9). With 6-mm simulated wall motion, the mean lesion volumes with MNS and MC were 60.2 ± 2.7 and 42.8 ± 8.4 mm(3), respectively, in the inferior catheter orientation (n = 11, p = <0.01*), 74.1 ± 5.8 and 54.2 ± 3.7 mm(3), respectively, in the superior catheter orientation (n = 10, p = <0.01*). During 6-mm simulated wall motion, the MC catheter and MNS catheter moved 5.2 ± 0.1 and 0 mm, respectively, in inferior orientation and 5.5 ± 0.1 and 0 mm, respectively, in the superior orientation on the ablation surface. CONCLUSIONS: The lesion dimensions were larger with MNS compared to MC in the presence of simulated wall motion, consistent with greater catheter stability. However, similar lesion dimensions were observed in the stationary model.

Role of endogenous endonucleases and tissue site in transfection and CpG-mediated immune activation after naked DNA injection.

DNA degradation is a fundamental problem for any gene therapy or genetic immunization approach, since destruction of incoming genes translates into loss of gene expression. To characterize the biology of DNA degradation after naked DNA injection, the location and levels of tissue nucleases were assessed. Extracts from the serum, kidney, and liver of mice had high levels of calcium-dependent endonuclease activity. High levels of acidic endonuclease activity were identified in the spleen, liver, kidney, and skin with little activity in skeletal or cardiac muscle. Relatively little exonuclease activity was observed in any tissue. The presence of endonucleases in the skin and muscle mediated degradation of 99% of naked DNA within 90 min of injection. This degradation most likely occurred in the extracellular space upstream of other cellular events. Despite this massive destruction, gross tissue nuclease levels did not determine skin-to-muscle transfection efficiency, or site-to-site transfection efficiency in the skin. While gross tissue nuclease levels do not appear to determine differences in transfection efficiency, the presence of robust tissue nuclease activity still necessitates that massive amounts of DNA be used to overcome the loss of 99% of expressible DNA. In addition to destroying genes, the nucleases may play a second role in genetic immunization by converting large plasmids into small oligonucleotides that can be taken up more easily by immune cells to stimulate CpG-dependent Th1 immune responses. For genetic immunization, vaccine outcome may depend on striking the right balance of nuclease effects to allow survival of sufficient DNA to express the antigen, while concomitantly generating sufficient amounts of immunostimulatory DNA fragments to drive Th1 booster effects. For gene therapy, all nuclease effects would appear to be negative, since these enzymes destroy gene expression while also stimulating cellular immune responses against transgene-modified host cells.

Afferent and efferent connections of the primary octaval nuclei in the clearnose skate, Raja eglanteria.

Horseradish peroxidase techniques were employed to trace the central projections of afferents from the individual endorgans of the membranous labyrinth and to delineate the efferent projections from the primary octaval nuclei to the spinal cord and midbrain octavolateralis area in the clearnose skate, Raja eglanteria. First-order octaval afferents project ipsilaterally to five primary octaval nuclei, namely: magnocellular, descending, posterior, anterior, and periventricular. Octaval afferents also terminate in the reticular formation, nucleus intermedius (primary mechanoreceptive lateral-line nucleus), and vestibulolateral lobe of the cerebellum. Each primary octaval nucleus receives afferent input from each labyrinthine endorgan, with the possible exception of macula neglecta input to the magnacellular nucleus. Within the anterior, descending, and to a lesser extent posterior and magnocellular nuclei, this input is largely nonoverlapping. Semicircular canal cristae afferents terminate ventrally, saccular and lagenar afferents dorsally, utricular afferents laterally, and macular neglecta afferents course ventrally but terminate largely dorsally within these nuclei. In the vestibulolateral lobe of the cerebellum, cristae afferents project primarily to the pars medialis, whereas macular endorgan afferents terminate in the pars lateralis. Primary afferent input to the reticular formation is predominantly from the horizontal canal crista. The densest projections to nucleus intermedius are from the utriculus and sacculus. Vestibulospinal projections originate primarily from the magnocellular and descending nuclei. Second-order auditory neurons are most likely located in dorsomedial parts of the descending and anterior nuclei. Cells in these nuclei project directly to the auditory area of the midbrain octavolateralis complex, but projections to this area originate predominantly from nuclei C1 and C2, which are possible superior olivary homologues.

Genetic immunization: applying recombinant DNA technology in the immune system to develop new vaccines.

Genetic immunization is a new method for vaccine development where the gene for an antigen is introduced into the host, such that the host cells produce vaccine antigens in vivo. By producing antigens intracellularly, genetic immunization is able to produce protective cellular immune responses as well as antibody responses to combat pathogens and cancer. Because genetic vaccines consist of plasmid DNA, the fundamental character of this vaccine can be manipulated at will by simple recombinant DNA techniques. This ability to easily modify and test novel vaccines combined with the power to drive both arms of the immune system, gives this technology a unique potential unavailable to other vaccines to combat the most difficult of disease targets. This review will discuss the current and future applications of this technology.