Pharmacokinetics and biodistribution of a collagen-targeted peptide amphiphile for cardiovascular applications.

Atherosclerosis remains a leading cause of death and disability around the world and a major driver of health care spending. Nanomaterials have gained widespread attention due to their promising potential for clinical translation and use. We have developed a collagen-targeted peptide amphiphile (PA)-based nanofiber for the prevention of neointimal hyperplasia after arterial injury. Our goal was to characterize the pharmacokinetics and biodistribution of the collagen-targeted PA to further its advancement into clinical trials. Collagen-targeted PA was injected into the internal jugular vein of Sprague Dawley rats. PA concentrations in plasma collected at various times after injection (0 to 72 hours) were measured by liquid chromatography-tandem mass spectrometry. Pharmacokinetics of the collagen-targeted PA were characterized by a three-compartment model, with an extremely rapid apparent elimination clearance resulting in a plasma concentration decrease of more than two orders of magnitude within the first hour after injection. This rapid initial decline in plasma concentration was not due to degradation by plasma components, as collagen-targeted PA was stable in plasma ex vivo for up to 3 hours. Indeed, cellular blood components appear to be partly responsible, as only 15% of collagen-targeted PA were recovered following incubation with whole blood. Nanofibers in whole blood also adhered to red blood cells (RBCs) and were engulfed by mononuclear cells. This work highlights the unique pharmacokinetics of our collagen-targeted PA, which differ from pharmacokinetics of small molecules. Because of their targeted nature, these nanomaterials should not require sustained elevated plasma concentrations to achieve a therapeutic effect the way small molecules typically do.

Development of Optimized Tissue-Factor-Targeted Peptide Amphiphile Nanofibers to Slow Noncompressible Torso Hemorrhage.

Noncompressible torso hemorrhage accounts for a significant portion of preventable trauma deaths. We report here on the development of injectable, targeted supramolecular nanotherapeutics based on peptide amphiphile (PA) molecules that are designed to target tissue factor (TF) and, therefore, selectively localize to sites of injury to slow hemorrhage. Eight TF-targeting sequences were identified, synthesized into PA molecules, coassembled with nontargeted backbone PA at various weight percentages, and characterized via circular dichroism spectroscopy, transmission electron microscopy, and X-ray scattering. Following intravenous injection in a rat liver hemorrhage model, two of these PA nanofiber coassemblies exhibited the most specific localization to the site of injury compared to controls (p < 0.05), as quantified using immunofluorescence imaging of injured liver and uninjured organs. To determine if the nanofibers were targeting TF in vivo, a mouse saphenous vein laser injury model was performed and showed that TF-targeted nanofibers colocalized with fibrin, demonstrating increased levels of nanofiber at TF-rich sites. Thromboelastograms obtained using samples of heparinized rat whole blood containing TF demonstrated that no clots were formed in the absence of TF-targeted nanofibers. Lastly, both PA nanofiber coassemblies decreased blood loss in comparison to sham and backbone nanofiber controls by 35-59% (p < 0.05). These data demonstrate an optimal TF-targeted nanofiber that localizes selectively to sites of injury and TF exposure, and, interestingly, reduces blood loss. This research represents a promising initial phase in the development of a TF-targeted injectable therapeutic to reduce preventable deaths from hemorrhage.

Development of Fractalkine-Targeted Nanofibers that Localize to Sites of Arterial Injury.

Atherosclerosis is the leading cause of death and disability around the world, with current treatments limited by neointimal hyperplasia. Our goal was to synthesize, characterize, and evaluate an injectable, targeted nanomaterial that will specifically bind to the site of arterial injury. Our target protein is fractalkine, a chemokine involved in both neointimal hyperplasia and atherosclerosis. We showed increased fractalkine staining in rat carotid arteries 24 h following arterial injury and in the aorta of low-density lipoprotein receptor knockout (LDLR-/-) mice fed a high-fat diet for 16 weeks. Three peptide amphiphiles (PAs) were synthesized: fractalkine-targeted, scrambled, and a backbone PA. PAs were ≥90% pure on liquid chromatography/mass spectrometry (LCMS) and showed nanofiber formation on transmission electron microscopy (TEM). Rats systemically injected with fractalkine-targeted nanofibers 24 h after carotid artery balloon injury exhibited a 4.2-fold increase in fluorescence in the injured artery compared to the scrambled nanofiber (p < 0.001). No localization was observed in the non-injured artery or with the backbone nanofiber. Fluorescence of the fractalkine-targeted nanofiber increased in a dose dependent manner and was observed for up to 48 h. These data demonstrate the presence of fractalkine after arterial injury and the localization of our fractalkine-targeted nanofiber to the site of injury and serve as the foundation to develop this technology further.

Targeted Nitric Oxide Delivery by Supramolecular Nanofibers for the Prevention of Restenosis After Arterial Injury.

AIMS: Cardiovascular interventions continue to fail as a result of arterial restenosis secondary to neointimal hyperplasia. We sought to develop and evaluate a systemically delivered nanostructure targeted to the site of arterial injury to prevent neointimal hyperplasia. Nanostructures were based on self-assembling biodegradable molecules known as peptide amphiphiles. The targeting motif was a collagen-binding peptide, and the therapeutic moiety was added by S-nitrosylation of cysteine residues. RESULTS: Structure of the nanofibers was characterized by transmission electron microscopy and small-angle X-ray scattering. S-nitrosylation was confirmed by mass spectrometry, and nitric oxide (NO) release was assessed electrochemically and by chemiluminescent detection. The balloon carotid artery injury model was performed on 10-week-old male Sprague-Dawley rats. Immediately after injury, nanofibers were administered systemically via tail vein injection. S-nitrosylated (S-nitrosyl [SNO])-targeted nanofibers significantly reduced neointimal hyperplasia 2 weeks and 7 months following balloon angioplasty, with no change in inflammation. INNOVATION: This is the first time that an S-nitrosothiol (RSNO)-based therapeutic was shown to have targeted local effects after systemic administration. This approach, combining supramolecular nanostructures with a therapeutic NO-based payload and a targeting moiety, overcomes the limitations of delivering NO to a site of interest, avoiding undesirable systemic side effects. CONCLUSION: We successfully synthesized and characterized an RSNO-based therapy that when administered systemically, targets directly to the site of vascular injury. By integrating therapeutic and targeting chemistries, these targeted SNO nanofibers provided durable inhibition of neointimal hyperplasia in vivo and show great potential as a platform to treat cardiovascular diseases.

Shape-Dependent Targeting of Injured Blood Vessels by Peptide Amphiphile Supramolecular Nanostructures.

Targeting of vascular intervention by systemically delivered supramolecular nanofibers after balloon angioplasty is described. Tracking of self-assembling peptide amphiphiles using fluorescence shows selective binding to the site of vascular intervention. Cylindrical nanostructures are observed to target the site of arterial injury, while spherical nanostructures with an equivalent diameter display no binding.

Surgical procedures before and after starting chronic hemodialysis in a predominantly male population with high prevalence of diabetes.

PURPOSE: The purposes of this study were to analyze changes in the frequency of different categories of surgical procedures after initiation of chronic hemodialysis (HD) and to identify the types of procedures associated with in-hospital postoperative mortality. METHODS: This was a retrospective analysis of surgical procedures performed in an incident HD population of 392 patients followed in the dialysis unit of one hospital over 15 years. Among these patients, 384 were men and 258 had diabetes mellitus. At the start of HD, age of the patients was 66.3 ± 11.2 years and Charlson index 5.35 ± 2.41. Rates of procedures per patient year (n/[pt-yr]), reported as mean (95% Confidence Interval [CI]), were compared by nonparametric methods. RESULTS: In the whole HD population, the overall rate of procedures increased in the HD period (pre-HD 0.125 [95% CI 0.101-0.149] n/[pt-yr]; HD 0.928 [95% CI 0.795-1.061] n/[pt-yr]; p<0.001). The increase, noted in patients with and without diabetes, reflected increases in the rates for both vascular access and non-vascular access procedures from the pre-HD to the HD period. Amputations and surgery for hip fractures accounted for the increase in the rates of procedures related to non-vascular access. Procedures associated with mortality in the HD period included amputations, hip repair and abdominal surgery for septic conditions. CONCLUSIONS: Rates of surgical procedures for vascular access, amputations, and hip fractures ?increased after the start of HD. Amputations and hip fractures, both potentially preventable, are associated with mortality in HD patients.

Hospitalizations before and after initiation of chronic hemodialysis.

Hospitalization rate is high in patients on chronic hemodialysis (HD). We investigated whether initiation of HD changes the rate and length of hospitalization. We analyzed hospitalizations in HD patients in one hospital over 15 years. We compared annual rate and length of hospitalizations, both presented as mean (95% confidence interval [CI]) between the pre-HD and HD period. Three hundred ninety-two patients, 98% men, 59% diabetic, and 66.3 ± 11.2 years old at the onset of HD, had 1016 hospitalizations in the pre-HD period (60.0 ± 42.9 months) and 1627 hospitalizations in the HD period (32.5 ± 25.9 months). Higher values were found in the HD than the pre-HD period for rate, (pre-HD 0.557 [95% CI 0.473-0.611], HD 2.198 [95% CI 1.997-2.399] admissions/[patient-year], P<0.001) and length (pre-HD 4.63 [95% CI 3.71-5.55], HD 28.07 [95% CI 23.55-32.59] days/patient-year], P<0.001) of hospitalizations for all causes, cardiac disease, infections, vascular access, peripheral vascular disease, metabolic disturbances, gastrointestinal diseases, and miscellaneous conditions, mainly respiratory illness and malignancy. Similar differences were found when we compared the year before and the year after the start of HD. Diabetics had higher all cause rate and length of hospitalizations than non-diabetics in the pre-HD and HD periods. The rate and length of hospitalizations was higher in the HD than the pre-HD period for both HD-specific conditions and conditions encountered in both HD and general populations. Study of factors specific to HD that may affect these conditions should constitute the first step toward improving the morbidity of patients on HD.

Hospital admissions in elderly patients on chronic hemodialysis.

BACKGROUND: The purpose of this study was to determine whether number of hospital admissions per patient per year (n/[pt-yr]) and hospital days per patient per year (d/[pt-yr]) differ between elderly and younger patients on chronic hemodialysis (HD). PATIENTS AND METHODS: In a retrospective cohort analysis of incident HD patients in one dialysis unit over 15 years, we compared 166 HD patients older than 70 years (77.1 ± 4.7 yrs) at the onset of HD (group A) and 216 patients younger than 70 years both at onset (57.1 ± 7.6 yrs) and at the end of the HD period (group B). Eighty (48.2%) of group A and 141 (65.3%) patients of group B had diabetes mellitus. RESULTS: No differences were noted in the overall hospitalization rate, presented as mean, {95% Confidence interval} (group A 2.40 {2.04-2.75}, group B 2.03 {1.89-2.16} n[pt-yr]) and days/[pt-year] (group A 33.6 {25.3-41.8}, group B 24.1 {18.9-29.23}). Group A had higher number of hospitalization days (P = 0.012) for surgery or trauma and higher rate (P = 0.045) and days (P = 0.041) of hospitalization for miscellaneous causes, primarily pulmonary disease, or malignancy. Among diabetic patients, group A had only a greater number of hospital days for cardiac disease (P = 0.050). Among patients without diabetes, group A had a higher number for hospital days for surgery or trauma (P = 0.027). All other univariate comparisons were not significant. Multiple linear regression identified comorbidity, quantified by the Charlson index, Caucasian race and poor compliance with the HD schedule as predictors of admission rate and days per year for vascular access issues and comorbidity, poor compliance, and advanced age at onset of HD as predictors of admission for causes other than vascular access related. CONCLUSION: Hospitalizations, which affect quality of life, differ little between elderly and younger patients on HD. Therefore, hospitalizations do not constitute an argument for restricting access to HD to elderly patients.

Hospitalizations in patients treated sequentially by chronic hemodialysis and continuous peritoneal dialysis.

It is not established whether hospitalizations are more frequent or longer in patients on peritoneal dialysis (PD) or chronic in-center hemodialysis (HD). Comorbidity is a major factor affecting the comparison of hospitalizations. To account for comorbidity, we compared hospitalizations between the PD and HD periods in 16 patients, 8 of whom were treated by PD first (group A), and 8, by HD first (group B). In group A, causes of renal failure were diabetes (n = 3), primary renal disease (n = 2), systemic disease (n = 2), and hereditary nephropathy (n = 1). Age at onset of PD was 53 +/- 11 years; duration of PD, 31 +/- 17 months; and duration of HD, 40 +/- 33 months. This group had 52 hospitalizations in the PD period and 80 hospitalizations in the HD period. Hospitalization rate (n/ patient-year) was 2.5 +/- 2.0 during PD and 3.0 +/- 3.0 during HD (nonsignificant), and duration of hospitalization (days/patient-year) was 19.6 +/- 15.5 during PD and 21.9 +/- 17.7 during HD (nonsignificant). The three most common causes of hospitalization were peritonitis (27%), other infections (21%), and cardiovascular disease (14%) in the PD period, and HD access problems (35%), infections (16%), and cardiovascular disease (12%) in the HD period. In group B, causes of renal failure were diabetes (n = 4), primary renal disease (n = 3), and hypertension (n = 1). Age at onset of HD was 56 +/- 10 years; duration of HD, 41 +/- 19 months; and duration of PD, 60 +/- 24 months. This group had 82 hospitalizations in the HD period and 76 hospitalizations in the PD period. Hospitalization rate was 3.0 +/- 2.4 during HD and 1.9 +/- 2.8 during PD (nonsignificant), and duration of hospitalization was 17.3 +/- 25.1 during HD and 12.7 +/- 21.3 during PD (nonsignificant). The three most common causes of hospitalization were HD access problems (40%), cardiovascular disease (19%), and infections (12%) in the HD period, and other infections (36%), cardiovascular disease (19%), and peritonitis (21%) in the PD period. In patients changing dialysis modalities, rate and duration of hospitalizations did not vary between HD and PD. The causes of hospitalization were similar in the HD and PD periods regardless of which modality was applied first.

Pulmonary function of herdsmen.

OBJECTIVE: To determine whether the pulmonary function deficit documented previously in Fulani children is also present in adult Fulani herdsmen in northern Nigeria. SUBJECTS AND METHODS: The subjects for this study consisted of adult Fulani men from the hamlet of Magama Gumau and adult non-Fulani men from the city of Jos. Age, height, weight, mid-arm circumference (MAC), triceps skin-fold thickness, forced vital capacity (FVC), forced expiratory volume in one second (FEV1), forced expiratory flow during the middle half of the FVC maneuver (FEF25-75%), and peak expiratory flow rate (PEF) were measured. Body mass index (BMI) and FEV1/FVC were calculated for all subjects. Multiple regression analysis was performed to identify correlations between pulmonary function parameters and anthropometric variables. RESULTS: The 44 Fulani subjects and 28 urban subjects were well-matched for age and height. The Fulani men weighed significantly less than the urban men (58.5+/-9.4 versus 67.4+/-11.3 kg, p <0.001) and consequently had significantly lower BMI, MAC, and triceps skin-fold thickness. The only significant difference in pulmonary function parameters between the two groups was in FEV1/FVC (0.93+/-0.1 versus 0.85+/-0.1, p <0.001). Small but significant correlations were found between pulmonary function parameters and anthropometric variables for both study populations. CONCLUSIONS: The pulmonary function deficits documented previously in Fulani children and adolescents were not present in adult Fulani men. However, the observed elevation in FEV1/FVC in the rural Fulani men as compared to their urban counterparts, which is often seen in restrictive pulmonary patterns, deserves further study.

Serum lipid profiles and risk of cardiovascular disease in three different male populations in northern Nigeria.

The Fulani of northern Nigeria are indigenous semi-nomadic pastoralists whose diet consists largely of dairy products. Despite their consumption of relatively large amounts of saturated fats, an earlier study showed that their total cholesterol, high-density lipoprotein (HDL), low-density lipoprotein (LDL) and serum triglyceride levels fall within the reference range of values for North Americans. Men in the cities of Jos and Abuja, two populations who also reside in northern Nigeria, differ from the Fulani with regard to diet and activity level. Males in both Jos and Abuja have diets consisting of high protein or carbohydrate and are more sedentary than the Fulani subjects. The main aims of the study were to measure the concentrations of various lipids in the blood serum of male urban dwellers in Jos and Abuja and to compare their blood lipid profiles with those of the rural Fulani (mean age 33.9 years). Blood serum samples from 118 men in Jos (mean age 37.9 years) and 77 men in Abuja (mean age 34.4 years) were analyzed for total cholesterol, triglycerides, LDL, HDL, homocysteine, folate, and vitamin B12. In addition to height and weight, systolic and diastolic blood pressures were measured. The mean total cholesterol, triglyceride, HDL and LDL values for the three groups of subjects fell within or close to the accepted range of values for North Americans. However, the Fulani males had HDL values (mean, 33.9 mg/dL) below the range of values prescribed for North Americans (>40 mg/dL). Moreover, the Fulani men and the men in Abuja had a total cholesterol/ HDL ratio of 4.2 and 4.0 respectively, which exceed the accepted value (< or =3.5) prescribed by the Columbia University. In all three populations, the incidence ofhomocysteinaemia (serum homocysteine > 12.4 micromol/L) was very high. Their mean homocysteine levels ranged from 14.7 to 16.7 pmol/L and could not be accounted for by folate or vitamin B12 status. The mean blood pressures of the Abuja (127/77 mm Hg) and the Fulani (120/74 mm Hg) men were within the normotensive range (<130/85 mm Hg). However, the mean blood pressures of the Jos males (131/85 mm Hg) indicated borderline hypertension. These data indicate that, with regard to serum lipids, urban and rural adult Nigerian males have generally favourable risk factors for cardiovascular disease when compared with healthy North Americans. All three sub-populations, however, have levels of homocysteine that are cause for concern vis-à-vis their overall health status.