A study of Chitosan and c-di-GMP as mucosal adjuvants for intranasal influenza H5N1 vaccine.

BACKGROUND: Highly pathogenic avian influenza A/H5N1 virus remains a potential pandemic threat, and it is essential to continue vaccine development against this subtype. A local mucosal immune response in the upper respiratory tract may stop influenza transmission. It is therefore important to develop effective intranasal pandemic influenza vaccines that induce mucosal immunity at the site of viral entry. OBJECTIVES: We evaluated the humoral and cellular immune responses of two promising mucosal adjuvants (Chitosan and c-di-GMP) for intranasal influenza H5N1 vaccine in a murine model. Furthermore, we evaluated the concept of co-adjuvanting an experimental adjuvant (c-di-GMP) with chitosan. METHODS: BALB/c mice were intranasally immunised with two doses of subunit NIBRG-14 (H5N1) vaccine (7·5, 1·5 or 0·3 µg haemagglutinin (HA) adjuvanted with chitosan (CSN), c-di-GMP or both adjuvants. RESULTS: All adjuvant formulations improved the serum and local antibody responses, with the highest responses observed in the 7·5 µg HA CSN and c-di-GMP-adjuvanted groups. The c-di-GMP provided dose sparing with protective single radial haemolysis (SRH), and haemagglutination inhibition (HI) antibody responses found in the 0·3 µg HA group. CSN elicited a Th2 response, whereas c-di-GMP induced higher frequencies of virus-specific CD4+T cells producing one or more Th1 cytokines (IFN-γ+, IL-2+, TNF-α+). A combination of the two adjuvants demonstrated effectiveness at 7·5 µg HA and triggered a more balanced Th cytokine profile. CONCLUSION: These data show that combining adjuvants can modulate the Th response and in combination with ongoing studies of adjuvanted intranasal vaccines will dictate the way forward for optimal mucosal influenza vaccines.

Chitosan-based delivery systems for mucosal vaccines.

INTRODUCTION: Mucosal vaccine development faces several challenges and opportunities. Critical issues for effective mucosal vaccination include the antigen-retention period that enables interaction with the lymphatic system, choice of adjuvant that is nontoxic and induces the required immune response and possibly an ability to mimic mucosal pathogens. Chitosan-based delivery systems are reviewed here as they address these issues and hence represent the most promising candidates for the delivery of mucosal vaccines. AREAS COVERED: A comprehensive literature search was conducted, to locate relevant studies published within the last 5 years. Mucosal delivery via nasal and oral routes is evaluated with respect to chitosan type, dosage forms, co-adjuvanting with novel adjuvants and modulation of the immune system. EXPERT OPINION: It is concluded that chitosan derivatives offer advantageous opportunities such as nanoparticle and surface charge manipulation that facilitate vaccine targeting. Nevertheless, these technologies represent a longer-term goal. By contrast, chitosan (unmodified form) with or without a co-adjuvant has significant toxicology and human data to support safe mucosal administration, and thus has the potential for earlier product introduction into the market.

The mucosal and systemic immune responses elicited by a chitosan-adjuvanted intranasal influenza H5N1 vaccine.

BACKGROUND: Development of influenza vaccines that induce mucosal immunity has been highlighted by the World Health Organisation as a priority (Vaccine 2005;23:1529). Dose-sparing strategies and an efficient mass-vaccination regime will be paramount to reduce the morbidity and mortality of a future H5N1 pandemic. OBJECTIVES: This study has investigated the immune response and the dose-sparing potential of a chitosan-adjuvanted intranasal H5N1 (RG-14) subunit (SU) vaccine in a mouse model. METHODS: Groups of mice were intranasally immunised once or twice with a chitosan (5 mg/ml)-adjuvanted SU vaccine [7·5, 15 or 30 µg haemagglutinin (HA)] or with a non-adjuvanted SU vaccine (30 µg HA). For comparison, another group of mice were intranasally immunised with a whole H5N1 (RG-14) virus (WV) vaccine (15 µg HA), and the control group consisted of unimmunised mice. RESULTS: The chitosan-adjuvanted SU vaccine induced an immune response superior to that of the non-adjuvanted SU vaccine. Compared with the non-adjuvanted SU group, the chitosan-adjuvanted SU vaccine elicited higher numbers of influenza-specific antibody-secreting cells (ASCs), higher concentrations of local and systemic antibodies and correspondingly an improved haemagglutination inhibition (HI) and single radial haemolysis (SRH) response against both the homologous vaccine strain and drifted H5 strains. We measured a mixed T-helper 1/T-helper 2 cytokine response in the chitosan-adjuvanted SU groups, and these groups had an increased percentage of virus-specific CD4(+) T cells producing two Thelper 1 (Th1) cytokines simultaneously compared with the non-adjuvanted SU group. Overall, the WV vaccine induced higher antibody concentrations in sera and an HI and SRH response similar to that of the chitosan-adjuvanted SU vaccine. Furthermore, the WV vaccine formulation showed a stronger bias towards a T-helper 1 profile than the SU vaccine and elicited the highest frequencies of CD4(+) Th1 cells simultaneously secreting three different cytokines (INFγ(+) , IL2(+) and INFα(+) ). As expected, two immunisations gave a better immune response than one in all groups. The control group had very low or not detectable results in the performed immunoassays. CONCLUSION: The cross-clade serum reactivity, improved B- and T-cell responses and dose-sparing potential of chitosan show that a chitosan-adjuvanted intranasal influenza vaccine is a promising candidate vaccine for further preclinical development.

Nasal vaccine innovation.

The current vaccine market is gaining momentum in the development of alternative administration routes namely intranasal, oral, topical, pulmonary, vaginal, and rectal; the nasal route offers the most promising opportunity for vaccine administration. It can enhance convenience, safety, elicit both local and systemic immune responses; thus potentially provide protection from pathogens at the site of entry. Nasal vaccine innovation comes with both opportunities and challenges. The innovative strategies used by industry and researchers to overcome the hurdles are discussed in this article: these include live-attenuated vaccines, adjuvants, mucoadhesives, particulate delivery systems, virus-like particles, vaccine manufacture, challenges of regulatory authorities, and the nasal vaccine impact on market potential. Critical issues for effective nasal vaccination are the antigen-retention period that enables its interaction with the lymphatic system and choice of an adjuvant that is nontoxic and induces the required immune response. Co-adjuvanting by means of a mucoadhesive technology addresses some of these issues. ChiSys(®), a natural bioadhesive with proven intranasal safety profile, has already demonstrated efficacy for several nasally delivered vaccines including norovirus. With the looming threat of a pandemic, alternatives such as intranasal vaccination will ultimately facilitate greater public compliance and rapid mass global vaccination.

Internal thoracic artery to left anterior descending anastomosis performed on the beating heart by endoscopic robot assistance using a new distal connector.

BACKGROUND: Automated distal connecting devices have been recently introduced to facilitate coronary anastomosis. This could have a large impact on the capacity of robotic systems to perform completely endoscopic off-pump bypass, where the quality of anastomosis and the prolonged operative time for the performance of the anastomosis have until now been cause for concern. Our group tried to determine the feasibility and efficacy of the JoMed distal graft connector using the ZEUS robotic system. METHODS: Six swine, with a mean weight of 25.8 (standard deviation [SD] 2.2) kg, underwent endoscopic off-pump internal thoracic artery-left anterior descending (ITA-LAD) anastomosis with a special stabilizing system using ZEUS robotic assistance. The anastomosis was performed with the JoMed distal connector. RESULTS: The connector was employed successfully in 4 of 6 cases using a special delivery instrument. Two animals fibrillated within 2 minutes after the application of proximal occluding snares and were excluded from the analysis. The total device deployment time was 2 minutes 4 seconds (SD 50 s) in 4 of 6 survivors, which remained hemodynamically stable and in sinus rhythm until euthanasia. Coronary angiography and transonic flow measurements were used to verify patency. CONCLUSION: The JoMed distal graft connector may facilitate the use of robot-assisted endoscopic bypass on a beating heart. Long-term patency issues will need to be assessed.

Nonoperative management of traumatic aortic injury.

BACKGROUND: The standard management for patients with blunt aortic injury is surgery; however, a small number of patients have been medically managed. The outcome of these nonoperatively managed patients is unknown. METHODS: Seven patients diagnosed as blunt aortic injury were managed without aortic surgery between January 1993 and April 2002, and their outcomes were retrospectively investigated. RESULTS: There were three men and four women, with a mean age of 48.7+/-22.7 years and Injury Severity Score of 37.7+/-16.9. The reason for nonoperative management was refusal of surgery (2), do-not-resuscitate order (1), diffuse brain injury (2), small intimal tear (1), and technical difficulty (1). Two patients died resulting from associated injuries. Five patients are alive, and in three patients complete resolution of aortic injury was observed. CONCLUSIONS: In selected patients with multiple associated injuries or severe comorbidity, nonoperative management after blunt aortic injury can be a treatment of choice.

Effect of chitosan on the intranasal absorption of salmon calcitonin in sheep.

The effects of a chitosan-based delivery system on the pharmacokinetics of intranasally administered salmon calcitonin (sCT) were investigated in a sheep model. In particular, the feasibility of producing a formulation with a comparable or improved bioavailability and/or less variability than the currently marketed nasal product (Miacalcin nasal spray, Novartis Pharmaceuticals) was assessed. A comparator (control) formulation comprising sCT solution was also tested. Sheep (n=6) were dosed intranasally according to a randomized crossover design. The intranasal sCT dose was 1100 IU (equivalent to approximately 17 IU kg-1). After completion of the nasal dosing legs, five of the sheep received 300 IU sCT (equivalent to approximately 5 IU kg-1) by subcutaneous injection to estimate relative bioavailability. After intranasal or subcutaneous dosing, serial blood samples were taken and plasma separated by centrifugation before measuring sCT concentrations by ELISA. Pharmacokinetic (non-compartmental) and statistical (analysis of variance or non-parametric alternative) analyses were performed. No systemic or local adverse effects were observed following intranasal or subcutaneous administration of sCT. The mean relative bioavailability of sCT from the chitosan solution was improved twofold compared with Miacalcin nasal spray and threefold compared with sCT control solution. Inter-animal variability in sCT absorption appeared to be lower with use of the chitosan-based solution compared with the control solution or commercial product. Based on the reported sheep data, a chitosan delivery system could offer the potential to significantly improve the intranasal absorption of sCT and reduce the variability in absorption. In the clinical setting, this may allow relatively lower doses of the drug to be given intranasally and/or lead to improvements in the efficacy or quality of intranasal therapy.

Intranasal delivery of recombinant human growth hormone (somatropin) in sheep using chitosan-based powder formulations.

The effectiveness of chitosan in promoting the intranasal bioavailability of recombinant human growth hormone (hGH) has been evaluated. hGH was formulated with chitosan to produce a powder blend (Formulation A) and granules (Formulation B) for intranasal administration. The in vivo pharmacokinetic performance of the formulations was evaluated in a group of six sheep in a randomised crossover study. A subcutaneous injection of hGH solution was administered as a control. The intranasal and subcutaneous doses of hGH were 0.3 and 0.03 mg/kg, respectively. The intranasal formulations appeared to be well tolerated. Mean bioavailabilities of hGH from Formulations A and B were 14 and 15%, respectively relative to subcutaneous injection. It is concluded that chitosan-based intranasal powder formulations may provide a practical means for non-injectable delivery of hGH and, potentially, other therapeutic protein molecules.

Intraoperative glucose control in diabetic and nondiabetic patients during cardiac surgery.

OBJECTIVE: The purpose of this study was to evaluate intraoperative glucose control. DESIGN: Prospective unblinded study. SETTING: Tertiary care center. PARTICIPANTS: Diabetic (n = 17) and nondiabetic (n = 23) patients undergoing elective cardiac surgery. INTERVENTIONS: Diabetics received a modified insulin regimen consisting of a fixed rate infusion of regular insulin, 10 U/m2/h, and a variable infusion of D10W, adjusted to maintain glucose between 101 to 140 mg/dL. MEASUREMENTS AND MAIN RESULTS: Baseline glucose was higher in diabetics versus nondiabetics (mean +/- standard error of the mean: 203 +/- 27 v 117 +/- 3 mg/dL, p < 0.005). After baseline, insulin levels were increased in diabetics to 410 to 568 microU/mL. Corresponding insulin levels in nondiabetics were 12 to 40 microU/mL. Compared with baseline, glucose was decreased by 10% +/- 29% in diabetics during hypothermic cardiopulmonary bypass and increased by 21% +/- 30% in nondiabetics (p < 0.005). After discontinuation of bypass, glucose was lower in diabetics (137 +/- 12 mg/dL) versus nondiabetics (162 +/- 8 mg/dL, p < 0.005). Nine diabetics had adequate intraoperative glycemic control during hypothermic bypass (glucose 123 +/- 8 mg/dL, insulin 550 +/- 68 microU/mL, glucose infusion rate 1.87 +/- 0.29 mg/kg/min), 6 approached adequate control near the end of surgery (glucose 147 +/- 8 mg/dL, insulin 483 +/- 86 microU/mL, glucose infusion rate 0.35 +/- 0.05 mg/kg/min), and 2 never achieved control. Diabetics with elevated initial glucose >300 mg/dL did not achieve adequate glycemic control. Four diabetics (3 with renal failure) required injection of 50% dextrose after bypass for hypoglycemia. CONCLUSION: Adequate glycemic control can be achieved in most diabetics during cardiac surgery using a modified insulin clamp technique provided initial glucose is <300 mg/dL.

Effective nasal influenza vaccine delivery using chitosan.

Nasal influenza vaccination may prove to be a good alternative to parenteral injection because of the enhancement of the mucosal immune response and the ease of vaccine administration. This study investigated the use of chitosan, a bioadhesive polymer, as a nasal delivery system with inactivated, subunit influenza vaccine. Subjects received nasally 15 or 7.5 microg of the standard inactivated trivalent influenza vaccine with chitosan or 15 microg of the same vaccine intramuscularly. Serum haemagglutination inhibition (HI) titres for all three vaccine components were measured prior to, and at time points up to 14 weeks after dosing. Serum HI titres following intranasal vaccination with the nasal chitosan-influenza vaccine met the criteria set by the Committee for Proprietary Medicinal Products in terms of seroprotection rate, seroconversion rate and mean fold increase of HI titre for at least one of the three antigens in the vaccination schedules used. These data show that nasal immunisation with chitosan plus trivalent inactivated influenza is a potentially effective, easily-administered form of vaccination.

Clinical diagnosis of actinic keratosis identifies an elderly population at high risk of developing skin cancer.

BACKGROUND: Development of actinic keratoses (AK) involves some of the same processes as nonmelanoma skin cancer and may serve as a marker for overall increased risk of skin cancer. OBJECTIVE: The objective of this study was to examine the risk of developing skin cancer in an elderly population with and without AK. METHODS: This was a retrospective observational study. Data from the 1992-1998 Medicare Current Beneficiary Study were used in the analyses. RESULTS: Multivariate analysis showed that the risk (odds ratio [OR]) of developing nonmelanoma or melanoma was increased more than sixfold (p < or = .0001) in patients with AK. An increased risk of skin cancer was found in whites (OR 4.3; p < or = .01) and increased age by year (OR 1.04; p < or = .01). Women were less likely to develop skin cancer (OR 0.58; p < or = .01). CONCLUSION: Using data from a nationally representive sample of the Medicare population, this study demonstrates that elders with AK are a population at high risk of developing cutaneous cancer.

Blunt injury of the innominate artery: a case report and review of literature.

Innominate artery injury after blunt trauma is uncommon and mostly observed at its origin from the aorta. We report here an unusual case of distal innominate artery injury associated with acute right subclavian occlusion. MEDLINE search of blunt traumatic injury to the innominate artery revealed a total of 132 case reports by the end of 2003, including this case report, and all these published studies were reviewed.

Redo-aortic valve replacement after previous bilateral internal thoracic artery bypass grafting.

BACKGROUND: Aortic valve replacement (AVR) after coronary artery bypass using bilateral internal thoracic arteries (ITAs) is a challenge. Management of these patent grafts and myocardial protection are important issues. Moreover the risk and outcome of these complex operations have not been clearly defined. METHODS: Eighteen consecutive patients (all male) who exhibited previous bilateral ITA grafts underwent subsequent AVR surgery from 1990-2001 at the Cleveland Clinic Foundation. Their medical records were retrospectively analyzed. RESULTS: At the time of reoperation, the mean age of the patients was 67 +/- 6.4 years and 33 out of 36 (92%) ITAs were patent. The interval between previous coronary bypass and aortic valve surgery was 10.3 +/- 5.3 years. All patients underwent redo-median sternotomy with aortic cannulation in 12 patients (67%) and femoral or axillary artery cannulation in 6 patients (33%). The patent ITAs were clamped during aortic cross-clamping in 15 patients. In 3 patients the ITAs were not dissected. These 3 patients underwent deep hypothermic arrest for myocardial protection. Concomitant coronary revascularization was performed in 8 patients (44%). There were no hospital deaths. One stroke occurred but there were no other major complications. Average intubation time was 23.1 +/- 27.1 hours, intensive care unit stay was 2.3 +/- 3.1 days, and postoperative hospital stay was 10.3 +/- 7.6 days. CONCLUSIONS: Reoperative aortic valve surgery in the patients with patent bilateral ITA grafts can be performed safely.

Robotically assisted endoscopic transmyocardial laser revascularization.

BACKGROUND: The surgical approach to transmyocardial laser revascularization (TMR) via thoracotomy has been approved for the treatment of angina in inoperable patients, but it has had limited use as a stand-alone procedure. OBJECTIVES: To assess the feasibility and efficacy of robotically assisted endoscopic TMR using the Zeus robotic surgical system (Computer Motion, USA) and an excimer TMR system (Spectranetics, USA) in a porcine model. METHODS: Five pigs weighing 20 kg to 40 kg were used. A 10 mm endoscope, two 5 mm instrument ports and a 1.4 mm TMR probe were introduced into the left hemithorax. Endoscopic TMR was performed on the anterolateral or lateral left ventricular wall using Zeus instruments to hold and guide the TMR probe. RESULTS: The mean system set-up time was 4.2+/-0.6 min, the mean port placement time was 11+/-1.7 min, the mean TMR procedure time was 5.0+/-0.7 min and the mean number of transmural TMR channels was 10+/-3.6. All pigs tolerated the entire procedure. Histological examination showed that transmural channels were successfully created. CONCLUSIONS: Endoscopic TMR using the Zeus system was feasible, in a pig model, in creating transmural laser channels on the anterior and lateral left ventricle, and may provide a significant advantage over the open chest approach in reducing perioperative morbidity and improving recovery.

Blunt injury of proximal innominate artery.

Innominate artery injury after blunt trauma is uncommon. We present a case of innominate artery injury, successfully treated with aorto-innominate bypass.

Intranasal immunization with genetically detoxified diphtheria toxin induces T cell responses in humans: enhancement of Th2 responses and toxin-neutralizing antibodies by formulation with chitosan.

We previously reported that intranasal immunization with a non-toxic mutant cross-reacting material (CRM)197 of diphtheria toxin, formulated with chitosan, generated protective neutralizing antibodies in mice and guinea pigs. Furthermore, we demonstrated that intranasal delivery of a powder formulation of the CRM197-based vaccine was well tolerated and significantly boosted antibody responses in adult volunteers. Here we report that intranasal booster immunization with CRM197 alone or with chitosan induced systemic T cell responses. We addressed for the first time the induction of T cell subtypes following intranasal vaccination in humans. Intranasal vaccination with CRM197, like parenteral immunization with a conventional diphtheria toxoid vaccine, enhanced antigen-specific IFN-gamma production. However, formulation of the nasal diphtheria vaccine with chitosan significantly augmented Th2-type responses, which correlated with protective levels of toxin-neutralizing antibodies in intranasally boosted individuals. The results suggest that vaccines capable of inducing strong Th2-type responses, such as CRM197 formulated with chitosan, have potential for the development of a protective mucosal vaccine against diphtheria in humans. Furthermore, our findings demonstrate that mucosal subunit vaccines with appropriate delivery systems have considerable potential for booster immunization of adults.

Migration of an inferior vena cava filter to the right ventricle and literature review.

Migration of an inferior vena cava (IVC) filter to the heart after placement is an extremely rare complication. The case of a 42-year-old man who presented with ventricular arrhythmia and tricuspid valve regurgitation, and underwent open heart surgery to extract an IVC filter from the right ventricle 12 days after infrarenal IVC filter placement, is reported.

Preservation of the anterior fat pad paradoxically decreases the incidence of postoperative atrial fibrillation in humans.

OBJECTIVES: The goal of this study was to determine if parasympathetic nerves in the anterior fat pad (FP) can be stimulated at the time of coronary artery bypass surgery (CABG), and if dissection of this FP decreases the incidence of postoperative atrial fibrillation (AF). BACKGROUND: The human anterior epicardial FP contains parasympathetic ganglia and is often dissected during CABG. Changes in parasympathetic tone influence the incidence of AF. METHODS: Fifty-five patients undergoing CABG were randomized to anterior FP preservation (group A) or dissection (group B). Nerve stimulation was applied to the FP before and after surgery. Sinus cycle length (CL) was measured during stimulation. The incidence of postoperative AF was recorded. RESULTS: Of the 55 patients enrolled, 26 patients were randomized to group A, and 29 patients were randomized to group B. In all of the 55 patients, the FP was identified before initiating cardiopulmonary bypass by CL prolongation with stimulation (865.5 +/- 147.9 ms vs. 957.9 +/- 155.1 ms, baseline vs. stimulation, p < 0.001). In group A, stimulation at the conclusion of surgery increased sinus CL (801.8 +/- 166.4 ms vs. 890.9 +/- 178.2 ms, baseline vs. stimulation, p < 0.001). In group B, repeat stimulation failed to increase sinus CL (853.6 +/- 201.6 ms vs. 841.4 +/- 198.4 ms, baseline vs. stimulation, p = NS). The incidence of postoperative AF in group A (7%) was significantly less than that in group B (37%) (p < 0.01). CONCLUSIONS: This is the first study demonstrating that direct stimulation of the human anterior epicardial FP slows sinus CL. This parasympathetic effect is eliminated with FP dissection. Preservation of the human anterior epicardial FP during CABG decreases incidence of postoperative AF.