Polysaccharide and monosaccharide guided liver delivery of Sorafenib Tosylate - A nano-strategic approach and comparative assessment of hepatospecificity.

Hepatospecific delivery by ligand based receptor targeting is an established strategy to augment therapy associated with liver diseases and disorders. Previously, we have investigated the effect of ligand headgroup on cellular uptake mediated by the asialoglycoprotein receptor by in silico and in vitro approach. In this paper, we report the design of agarose based liposomes for delivery to liver cancer cells and provide a proof of concept of the targeting efficiency against galactose liposomes using an in vivo approach. Sorafenib Tosylate loaded targeting liposomes were developed and optimized using factorial design. Comparative evaluation including cell cytotoxicity, pharmacokinetics and biodistribution and hepatospecific uptake was performed for both the liposomal systems. The formulations possessed a particle size of 150 - 180 nm and a zeta potential of 30 - 60 mV depending on the amount of ligand and drug loading, with more than 90% entrapment efficiency. A two-fold increase in cytotoxicity was observed with agarose-based liposomes as compared to galactose based liposomes. In vivo PK evaluation indicated a reduction in half life of drug when loaded in agarose ligand loaded system, probably due to greater uptake in the liver as evidenced in biodistribution study. Intrahepatic disposition revealed a higher PC/NPC uptake ratio with the targeted systems as compared to conventional liposomes, although the agarose-based system resulted in highest uptake ratio. A biocompatible platform for specific delivery of drugs to hepatocytes was established validating a rational approach to design liver targeting systems.

Polysaccharide conjugates surpass monosaccharide ligands in hepatospecific targeting - Synthesis and comparative in silico and in vitro assessment.

Ligands with the polysaccharide headgroups have been recently reported by our group to possess enhanced interaction with asialoglycoprotein receptor (ASGPR) in silico as compared to ligands having galactose moieties. This enhanced interaction is a result of the polymer's backbone support in anchoring the ligand in a specific orientation within the bilayer. In this paper, we have attempted to provide an in vitro proof of concept by performing a comparative evaluation of polysaccharide and monosaccharide-based ligands. Docking was performed to understand interaction with ASGPR in silico. Agarose and galactose conjugates with behenic acid were synthesized, purified, and characterized to yield biocompatible hepatospecific ligands which were incorporated into nanoliposomes. Cellular internalization of these targeted liposomes was studied using confocal microscopy and flow cytometry. The toxicity potential was assessed in vivo. Results indicated that the polysaccharide-based ligand increased cellular uptake due to better interaction with the receptor as compared to ligand bearing a single galactose group. In addition to developing novel liver targeting ligands, the study also established proof of concept that has been suggested by earlier in silico investigations. The approach can be used to design targeting ligands and develop formulations with improved targeting efficacy.

Lipid nanoconstructs for superior hepatoprotection: In vitro assessments as predictive tool for in vivo translation.

AIM: To investigate comparative in vitro and in vivo performance of lipid vesicular and particulate systems in escalating oral bioavailability for superior hepatoprotection. MATERIALS AND METHODS: Systems were fabricated using easy to scale up process and novel excipients to deliver Silibinin. In vitro characterization followed by pharmacokinetic and pharmacodynamic evaluation in rats was conducted to establish a correlation. RESULTS: Nanoformulations resulted in 20 fold increase in solubilisation and significant increase in permeation. 2.5 fold increase in bioavailability was evident in vivo. Vesicles demonstrated greatest hepatoprotective potential in efficacy study. CONCLUSION: The findings establish a link between in vitro and in vivo performance to rank order lipid nanoartchitects. Concurrently, a significant potential in therapeutic intervention of hepatotoxicity is envisaged as elucidated.

Using tongue depressors to aid cord localization during collagenase injection for Dupuytren disease.

The aim of this paper was to introduce a simple and effective method to aid in isolation and stabilization of Dupuytren cords for collagenase injection. Tongue depressors were used to isolate and stabilize the cord during the injection procedure. The area to be injected was sterilely prepared. An assistant was then directed to place a tongue depressor on both sides of the cord. A total of 35 patients with Dupuytren disease were treated. Follow-up lasted two years. Post-operative Disabilities of the Arm, Shoulder and Hand (DASH) scores was 4±2. Health-related quality of life measured with the EQ-5D index was 0.89±0.4. Recurrence rates of metacarpophalangeal joint and proximal interphalangeal joint were 11% and 14% respectively, using a flexion contracture of 20° to define recurrence. Collagenase treatment using a modified injection method with the aid of tongue depressors are a safe, effective way to treat Dupuytren contractures of the fingers. The technique can isolate the cord, which improves visualization of the cord. It may allow improved accuracy with needle placement and helps to decrease the complications and recurrence.

Cationic nanoemulsions as potential carriers for intracellular delivery.

Successful cytosolic delivery enables opportunities for improved treatment of various genetic disorders, infectious diseases and cancer. Cationic nanoemulsions were designed using alternative excipients and evaluated for particle size, charge, effect of sterilization on its stability, DNA condensation potential and cellular uptake efficiency. Various concentrations of non-ionic and ionic stabilizers were evaluated to design formula for colloidally stable cationic nanoemulsion. The nanoemulsion comprised of 5% Capmul MCM, 0.5% didodecyldimethylammonium bromide (DDAB), 1% phospholipid, 1% Poloxamer 188 and 2.25% glycerol and possessed particle size of 81.6 ± 3.56 nm and 137.1 ± 1.57 nm before and after steam sterilization, respectively. DNA condensation studies were carried out at various nanoemulsion: DNA ratios ranging from 1:1 to 10:1. Cell uptake studies were conducted on human embryonic kidney (HEK) cell lines which are widely reported for transfection studies. The nanoemulsions showed excellent cellular uptake as evaluated by fluorescence microscopy and flow cytometry. Overall, a colloidally stable cationic nanoemulsion with good DNA condensation ability was successfully fabricated for efficient cytosolic delivery and potential for in vivo effectiveness.

Transcranial sonography and functional imaging in glucocerebrosidase mutation Parkinson disease.

BACKGROUND: Heterozygous glucocerebrosidase (GBA) mutations are the leading genetic risk factor for Parkinson disease, yet imaging correlates, particularly transcranial sonography, have not been extensively described. METHODS: To determine whether GBA mutation heterozygotes with Parkinson disease demonstrate hyperechogenicity of the substantia nigra, transcranial sonography was performed in Ashkenazi Jewish Parkinson disease subjects, tested for the eight most common Gaucher disease mutations and the LRRK2 G2019S mutation, and in controls. [(18)F]-fluorodeoxyglucose or [(18)F]-fluorodopa positron emission tomography is also reported from a subset of Parkinson disease subjects with heterozygous GBA mutations. RESULTS: Parkinson disease subjects with heterozygous GBA mutations (n = 23) had a greater median maximal area of substantia nigral echogenicity compared to controls (n = 34, aSNmax = 0.30 vs. 0.18, p = 0.007). There was no difference in median maximal area of nigral echogenicity between Parkinson disease groups defined by GBA and LRRK2 genotype: GBA heterozygotes; GBA homozygotes/compound heterozygotes (n = 4, aSNmax = 0.27); subjects without LRRK2 or GBA mutations (n = 32, aSNmax = 0.27); LRRK2 heterozygotes/homozygotes without GBA mutations (n = 27, aSNmax = 0.28); and GBA heterozygotes/LRRK2 heterozygotes (n = 4, aSNmax = 0.32, overall p = 0.63). In secondary analyses among Parkinson disease subjects with GBA mutations, maximal area of nigral echogenicity did not differ based on GBA mutation severity or mutation number. [(18)F]-fluorodeoxyglucose (n = 3) and [(18)F]-fluorodopa (n = 2) positron emission tomography in Parkinson disease subjects with heterozygous GBA mutations was consistent with findings in idiopathic Parkinson disease. CONCLUSIONS: Both transcranial sonography and positron emission tomography are abnormal in GBA mutation associated Parkinson disease, similar to other Parkinson disease subjects.

Network correlates of disease severity in multiple system atrophy.

OBJECTIVE: Multiple system atrophy (MSA), the most common of the atypical parkinsonian disorders, is characterized by the presence of an abnormal spatial covariance pattern in resting state metabolic brain images from patients with this disease. Nonetheless, the potential utility of this pattern as a MSA biomarker is contingent upon its specificity for this disorder and its relationship to clinical disability in individual patients. METHODS: We used [(18)F]fluorodeoxyglucose PET to study 33 patients with MSA, 20 age- and severity-matched patients with idiopathic Parkinson disease (PD), and 15 healthy volunteers. For each subject, we computed the expression of the previously characterized metabolic covariance patterns for MSA and PD (termed MSARP and PDRP, respectively) on a prospective single-case basis. The resulting network values for the individual patients were correlated with clinical motor ratings and disease duration. RESULTS: In the MSA group, disease-related pattern (MSARP) values were elevated relative to the control and PD groups (p < 0.001 for both comparisons). In this group, MSARP values correlated with clinical ratings of motor disability (r = 0.57, p = 0.0008) and with disease duration (r = -0.376, p = 0.03). By contrast, MSARP expression in the PD group did not differ from control values (p = 1.0). In this group, motor ratings correlated with PDRP (r = 0.60, p = 0.006) but not with MSARP values (p = 0.88). CONCLUSIONS: MSA is associated with elevated expression of a specific disease-related metabolic pattern. Moreover, differences in the expression of this pattern in patients with MSA correlate with clinical disability. The findings suggest that the MSARP may be a useful biomarker in trials of new therapies for this disorder.

Network correlates of the cognitive response to levodopa in Parkinson disease.

OBJECTIVE: Cognitive dysfunction is common in Parkinson disease (PD), even early in its clinical course. This disease manifestation has been associated with impaired verbal learning performance as well as abnormal expression of a specific PD-related cognitive spatial covariance pattern (PDCP). It is not known, however, how this metabolic network relates to the cognitive response to dopaminergic therapy on the individual patient level. METHODS: We assessed treatment-mediated changes in verbal learning and PDCP expression in 17 patients with PD without dementia who underwent cognitive testing and metabolic imaging in the unmedicated and levodopa-treated conditions. We also determined whether analogous changes were present in 12 other patients with PD without dementia who were evaluated before and during the treatment of cognitive symptoms with placebo. RESULTS: Levodopa-mediated changes in verbal learning correlated with concurrent changes in PDCP expression (r = -0.60, p < 0.01). The subset of patients with meaningful cognitive improvement on levodopa (n = 8) exhibited concurrent reductions in PDCP expression (p < 0.01) with treatment; network modulation was not evident in the remaining subjects. Notably, the levodopa cognitive response correlated with baseline PDCP levels (r = 0.70, p = 0.002). By contrast, placebo did not affect PDCP expression, even in the subjects (n = 7) with improved verbal learning during treatment. CONCLUSIONS: These findings suggest that cognitive dysfunction in PD may respond to treatment depending upon the degree of baseline PDCP expression. Quantification of treatment-mediated network changes can provide objective information concerning the efficacy of new agents directed at the cognitive manifestations of this disease.

Abnormal metabolic brain networks in Tourette syndrome.

OBJECTIVES: To identify metabolic brain networks that are associated with Tourette syndrome (TS) and comorbid obsessive-compulsive disorder (OCD). METHODS: We utilized [(18)F]-fluorodeoxyglucose and PET imaging to examine brain metabolism in 12 unmedicated patients with TS and 12 age-matched controls. We utilized a spatial covariance analysis to identify 2 disease-related metabolic brain networks, one associated with TS in general (distinguishing TS subjects from controls), and another correlating with OCD severity (within the TS group alone). RESULTS: Analysis of the combined group of patients with TS and healthy subjects revealed an abnormal spatial covariance pattern that completely separated patients from controls (p < 0.0001). This TS-related pattern (TSRP) was characterized by reduced resting metabolic activity of the striatum and orbitofrontal cortex associated with relative increases in premotor cortex and cerebellum. Analysis of the TS cohort alone revealed the presence of a second metabolic pattern that correlated with OCD in these patients. This OCD-related pattern (OCDRP) was characterized by reduced activity of the anterior cingulate and dorsolateral prefrontal cortical regions associated with relative increases in primary motor cortex and precuneus. Subject expression of OCDRP correlated with the severity of this symptom (r = 0.79, p < 0.005). CONCLUSION: These findings suggest that the different clinical manifestations of TS are associated with the expression of 2 distinct abnormal metabolic brain networks. These, and potentially other disease-related spatial covariance patterns, may prove useful as biomarkers for assessing responses to new therapies for TS and related comorbidities.

Transcriptional expression and gelatinolytic activity of matrix metalloproteinases in Henoch-Schonlein purpura.

AIM: Accelerated extracellular matrix breakdown caused by the increased activity of matrix metalloproteinases (MMPs) has been implicated in several rheumatological disorders and systemic vasculitides, especially Takayasu's arteritis and Kawasaki disease. Therefore, the aim of the present study was to investigate the potential role of MMPs in Henoch-Schonlein purpura (HSP), an acute type of systemic vasculitis in children. METHODS: We studied the activity of MMP-2 and MMP-9 in the sera using gelatin zymography and the transcriptional expression in peripheral blood mononuclear cells using semi-quantitative RT-PCR in 20 patients with HSP in acute and convalescent phase and in 20 healthy children, who were siblings of the subjects with same age group. RESULTS: All 20 children with HSP showed increased levels of serum activity of MMP-2 and MMP-9 in acute phase as compared with their convalescent phase [MMP-2 (p > 0.05); MMP-9 (p > 0.05)] and their control counterparts [MMP-2 (p < 0.001); MMP-9 (p < 0.001)]. Similarly, transcriptional expression of MMPs was found to be higher in the acute phase of HSP than in convalescent phase [MMP-2 (p < 0.05); MMP-9 (p < 0.001)] and in their healthy controls [MMP-2 (p < 0.001); MMP-9 (p < 0.01)]. CONCLUSION: The presence of excessive transcriptional expression and gelatinolytic activity of MMPs may be downstream to the actual aetiopathogenetic factors.

Abnormal striatal and thalamic dopamine neurotransmission: Genotype-related features of dystonia.

OBJECTIVE: To determine whether changes in D(2) receptor availability are present in carriers of genetic mutations for primary dystonia. METHODS: Manifesting and nonmanifesting carriers of the DYT1 and DYT6 dystonia mutations were scanned with [(11)C] raclopride (RAC) and PET. Measures of D(2) receptor availability in the caudate nucleus and putamen were determined using an automated region-of-interest approach. Values from mutation carriers and healthy controls were compared using analysis of variance to assess the effects of genotype and phenotype. Additionally, voxel-based whole brain searches were conducted to detect group differences in extrastriatal regions. RESULTS: Significant reductions in caudate and putamen D(2) receptor availability were evident in both groups of mutation carriers relative to healthy controls (p < 0.001). The changes were greater in DYT6 relative to DYT1 carriers (-38.0 +/- 3.0% vs -15.0 +/- 3.0%, p < 0.001). By contrast, there was no significant difference between manifesting and nonmanifesting carriers of either genotype. Voxel-based analysis confirmed these findings and additionally revealed reduced RAC binding in the ventrolateral thalamus of both groups of mutation carriers. As in the striatum, the thalamic binding reductions were more pronounced in DYT6 carriers and were not influenced by the presence of clinical manifestations. CONCLUSIONS: Reduced D(2) receptor availability in carriers of dystonia genes is compatible with dysfunction or loss of D(2)-bearing neurons, increased synaptic dopamine levels, or both. These changes, which may be present to different degrees in the DYT1 and DYT6 genotypes, are likely to represent susceptibility factors for the development of clinical manifestations in mutation carriers.

Metabolic abnormalities associated with mild cognitive impairment in Parkinson disease.

OBJECTIVE: To use (18)F-fluorodeoxyglucose (FDG) and PET to investigate changes in regional metabolism associated with mild cognitive impairment (MCI) in Parkinson disease (PD). Cognitive abnormalities are common in PD. However, little is known about the functional abnormalities that underlie the manifestations of MCI in this disorder. METHODS: We used FDG PET to measure regional glucose metabolism in patients with PD with multiple-domain MCI (MD-MCI; n = 18), with single-domain MCI (SD-MCI; n = 15), and without MCI (N-MCI; n = 18). These patients were matched for age, education, disease duration, and motor disability. Maps of regional metabolism in the three groups were compared using statistical parametric mapping (SPM). We also computed the expression of a previously validated cognition-related spatial covariance pattern (PDCP) in the patient groups and in an age-matched healthy control cohort (n = 15). PDCP expression was compared across groups using analysis of variance. RESULTS: SPM revealed decreased prefrontal and parietal metabolism (p < 0.001) in MD-MCI relative to N-MCI, as well as an increase in brainstem/cerebellar metabolism (p < 0.001) in this group. In these regions, SD-MCI occupied an intermediate position between the two other groups. PDCP expression was abnormally elevated in the N-, SD-, and MD-MCI groups (p < 0.05), increasing stepwise with worsening cognitive impairment (p < 0.01). CONCLUSIONS: Early cognitive decline in Parkinson disease as defined by mild cognitive impairment is associated with discrete regional changes and abnormal metabolic network activity. The quantification of these alterations with (18)F-fluorodeoxyglucose PET may allow for the objective assessment of the progression and treatment of this disease manifestation.

'Induction of inflammatory gene expression by THP-1 macrophages cultured in normocholesterolaemic hypertensive sera and modulatory effects of green tea polyphenols'.

Hypertension is a disorder controlled by multiple genes and inflammation and vascular remodelling of arteries have been implicated in pathogenesis of this disease. Green tea polyphenols (GrTPs) are rich in antioxidants and are known to inhibit inflammatory responses. A significant time-dependent increase in mRNA expression of both IL-6 and MMP-9 were observed in THP-1 macrophages when cultured in normocholesterolaemic hypertensive sera (P<0.05).

Thalamic metabolism and symptom onset in preclinical Huntington's disease.

The neural basis for the transition from preclinical to symptomatic Huntington's disease (HD) is unknown. We used serial positron emission tomography (PET) imaging in preclinical HD gene carriers (p-HD) to assess the metabolic changes that occur during this period. Twelve p-HD subjects were followed longitudinally with [11C]-raclopride and [18F]-fluorodeoxyglucose PET imaging, with scans at baseline, 18 and 44 months. Progressive declines in striatal D2-receptor binding were correlated with concurrent changes in regional metabolism and in the activity of an HD-related metabolic network. We found that striatal D2 binding declined over time (P < 0.005). The activity of a reproducible HD-related metabolic covariance pattern increased between baseline and 18 months (P < 0.003) but declined at 44 months (P < 0.04). These network changes coincided with progressive declines in striatal and thalamic metabolic activity (P < 0.01). Striatal metabolism was abnormally low at all time points (P < 0.005). By contrast, thalamic metabolism was elevated at baseline (P < 0.01), but fell to subnormal levels in the p-HD subjects who developed symptoms. These findings were confirmed with an MRI-based atrophy correction for each individual PET scan. Increases in network expression and thalamic glucose metabolism may be compensatory for early neuronal losses in p-HD. Declines in these measures may herald the onset of symptoms in gene carriers.

Genetic aspects of restless legs syndrome.

Restless legs syndrome (RLS), also known as Ekbom syndrome, is a common movement disorder with sensorimotor symptoms occurring during sleep and quiet wakefulness. The underlying cause for RLS is unknown but genetic influences play a strong part in the pathogenesis of RLS, particularly when the condition starts at a young age. This review explores the genetic basis of RLS and related phenotypic variations. Recently, three loci showing vulnerability to RLS have been described in French-Canadian and Italian families in chromosomes 12q, 14q and 9q, emphasising on an autosomal dominant mode of inheritance. These have been labelled RLS1, RLS2 and RLS3, respectively. However, specific causative mutations remain elusive and no linkage analysis has been identified so far in the candidate genes investigated in RLS.

The range and nature of sleep dysfunction in untreated Parkinson's disease (PD). A comparative controlled clinical study using the Parkinson's disease sleep scale and selective polysomnography.

In this study we have explored the nature and range of sleep dysfunction that occurs in untreated Parkinson's disease (PD) comparing data obtained from the use of the Parkinson's disease sleep scale (PDSS) in an untreated PD patient group compared to advanced PD and healthy controls. 25 untreated (drug-naive, DNPD) PD patients (mean age 66.9 years, range 53-80, 18 males) completed the validated Parkinson's disease sleep scale (PDSS), mean duration of PD was 2.1 years (1-10, up to 4 years in all except one patient with tremulous PD reporting tremor duration of 10 years) and mean Hoehn and Yahr score 1.9 (1-3). Data were compared to 34 advanced PD (mean age 70.2 years, range 51-88, 23 male), mean duration of PD 11 years (range 4-22), mean Hoehn and Yahr score 3.4 (3-5) and PDSS data obtained from 131 healthy controls (mean age 66.6 years, range 50-93, 56 males). Total PDSS scores and PDSS sub-items, except PDSS item 2, were highly significantly different (p<0.001) between DNPD, advanced PD and controls. Controls reported higher mean PDSS scores than both groups of patients, and advanced cases reported lower (mean+/-S.D.) PDSS scores (86.95+/-20.78) than drug-naive (105.72+/-21.5) (p<0.001). Logistic regression analysis showed that items PDSS8 (nocturia), PDSS11 (cramps), PDSS12 (dystonia), PDSS13 (tremor), and PDSS15 (daytime somnolence) were significantly impaired in DNPD compared to controls while PDSS7 (nighttime hallucinations) additionally separated advanced PD from DNPD. In a subgroup of 11 advanced PD cases (mean age 62 years, range=49-84 years, mean Hoehn and Yahr score 2.5, range=1-3) with high Epworth Sleepiness Scale (ESS) scores (mean 14.5), low item 15 PDSS score (mean 4.7) and complaints of severe daytime sleepiness, underwent detailed overnight polysomnography (PSG) studies, all showing abnormal sleep patterns. We conclude that nocturia, nighttime cramps, dystonia, tremor and daytime somnolence seem to be the important nocturnal disabilities in DNPD and some of these symptoms may be reminiscent of "off" period related symptoms even though patients are untreated. Furthermore, polysomnography in "sleepy" PD patients may help diagnose unrecognised conditions such as periodic limb movement of sleep (PLMS), obstructive sleep apnoea (OSA) and REM Sleep Behaviour Disorder.

Sleep-related problems of Parkinson's disease.

OBJECTIVE: To define the epidemiology, characteristics and aetiology of nocturnal symptoms and sleep disorders in patients with Parkinson's disease (PD) and evaluate the available methods for their diagnosis and management. METHODS: A review of the English-language literature pertaining to sleep disturbances associated with PD, using the Medline database and bibliographies in relevant articles. RESULTS: Sleep-related problems specific to PD may occur early and even predate the diagnosis of the disease but are generally more frequent and more severe in patients with advanced PD. These problems can seriously compromise patients' quality of life and lead to impaired functioning in daily activities. Scales designed specifically for the assessment of sleep problems in patients with PD have recently been developed. Evidence base for the treatment of sleep disturbances in PD is poor, and only nocturnal akinesia, excessive day-time sleepiness and rapid eye movement behaviour disorder have been partially addressed. CONCLUSIONS: Sleep disorders associated with PD are a common and under-recognised problem. The assessment of sleep should be part of the routine evaluation of patients with PD, and large-scale controlled therapeutic trials are necessary.