Systemic dendrimer-drug nanomedicines for long-term treatment of mild-moderate cerebral palsy in a rabbit model.

BACKGROUND: Neuroinflammation mediated by microglia plays a central role in the pathogenesis of perinatal/neonatal brain injury, including cerebral palsy (CP). Therapeutics mitigating neuroinflammation potentially provide an effective strategy to slow the disease progression and rescue normal brain development. Building on our prior results which showed that a generation-4 hydroxyl poly(amidoamine) (PAMAM) dendrimer could deliver drugs specifically to activated glia from systemic circulation, we evaluated the sustained efficacy of a generation-6 (G6) hydroxyl-terminated PAMAM dendrimer that showed a longer blood circulation time and increased brain accumulation. N-acetyl-L-cysteine (NAC), an antioxidant and anti-inflammatory agent that has high plasma protein binding properties and poor brain penetration, was conjugated to G6-PAMAM dendrimer-NAC (G6D-NAC). The efficacy of microglia-targeted G6D-NAC conjugate was evaluated in a clinically relevant rabbit model of CP, with a mild/moderate CP phenotype to provide a longer survival of untreated CP kits, enabling the assessment of sustained efficacy over 15 days of life. METHODS: G6D-NAC was conjugated and characterized. Cytotoxicity and anti-inflammatory assays were performed in BV-2 microglial cells. The efficacy of G6D-NAC was evaluated in a rabbit model of CP. CP kits were randomly divided into 5 groups on postnatal day 1 (PND1) and received an intravenous injection of a single dose of PBS, or G6D-NAC (2 or 5 mg/kg), or NAC (2 or 5 mg/kg). Neurobehavioral tests, microglia morphology, and neuroinflammation were evaluated at postnatal day 5 (PND5) and day 15 (PND15). RESULTS: A single dose of systemic 'long circulating' G6D-NAC showed a significant penetration across the impaired blood-brain-barrier (BBB), delivered NAC specifically to activated microglia, and significantly reduced microglia-mediated neuroinflammation in both the cortex and cerebellum white matter areas. Moreover, G6D-NAC treatment significantly improved neonatal rabbit survival rate and rescued motor function to nearly healthy control levels at least up to 15 days after birth (PND15), while CP kits treated with free NAC died before PND9. CONCLUSIONS: Targeted delivery of therapeutics to activated microglia in neonatal brain injury can ameliorate pro-inflammatory microglial responses to injury, promote survival rate, and improve neurological outcomes that can be sustained for a long period. Appropriate manipulation of activated microglia enabled by G6D-NAC can impact the injury significantly beyond inflammation.

Assessment of salivary total antioxidant levels and oral health status in children with cerebral palsy.

UNLABELLED: Oxidative stress plays a pivotal role in the pathogenesis of neurological disorders. Saliva may constitute a first line of defence, against free radical-modified oxidative stress. The objective of the present study was to evaluate Total Antioxidant Capacity (TAC), levels of Nitric Oxide (NO), and Sialic Acid (SA) in saliva of cerebral palsied children. STUDY DESIGN: Thirty four non-institutionalized children in the age group of 7-12 years having cerebral palsy formed the study group. The control group consisted of thirty three normal, healthy children. The W.H.O. criteria was used for diagnosis and recording of dental caries. Oral hygiene status was assessed using the Simplified Oral Hygiene Index OHI-S. Estimation of Total Antioxidant Capacity, levels of Nitric Oxide and Sialic Acid in saliva was done. Data obtained was subjected to statistical analysis. RESULTS: Children with CP had higher deft scores than that of normal children. Oral hygiene of children with CP was significantly poorer than that of normal children. Total Antioxidant Capacity of saliva was significantly higher in normal children than cerebral palsied children. Levels of Sialic Acid in saliva were significantly higher in cerebral palsy children. CONCLUSIONS: In children with CP, TAC of saliva showed an inverse relation with dental caries.

Increased salivary immunoglobulin A and reduced α-amylase activity in whole saliva from spastic cerebral palsy individuals.

BACKGROUND: Salivary immunoglobulin A (SIgA) together with innate defenses such as α-amylase, provides the 'first line of defense' against pathogens present at mucosal surfaces. This study aimed to evaluate salivary α-amylase and immunoglobulin A (IgA) in whole saliva of spastic cerebral palsy (CP) individuals. METHODS: Whole saliva was collected from 22 CP and 24 sibling volunteers with no neurological damage control groups (CG) (aged 7-14 years). The salivary flow rate, total protein and SIgA concentrations, and α-amylase activity were determined. RESULTS: The CP group presented higher salivary flow rate (35%) and lower total protein concentration (18%) compared with the CG (P ≤ 0.05). CPG had higher absolute (68%, µg SIgA/ml) and relative (55%, µg SIgA/mg prot and 108%, µg SIgA/min) concentrations of IgA compared with the CG (P ≤ 0.05). CPG had lower relative α-amylase activity (15% mg malt/mg prot and 33%, mg malt/min) compared with the CG (P ≤ 0.05). CONCLUSION: This study concluded that CP individuals presented alterations in the profile of salivary proteins involved in the defense system of the oral cavity.

The Impact of Oral-Gut Inflammation in Cerebral Palsy.

Background: Oral-gut inflammation has an impact on overall health, placing subjects at risk to acquire chronic conditions and infections. Due to neuromotor disturbances, and medication intake, cerebral palsy (CP) subjects present intestinal constipation, impacting their quality of life (QOL). We aimed to investigate how oral inflammatory levels predicted gut phenotypes and response to therapy. Methods: A total of 93 subjects aging from 5 to 17 years were included in the study, and assigned into one of the 4 groups: CP with constipation (G1, n = 30), CP without constipation (G2, n = 33), and controls without CP with constipation (G3, n = 07) and without CP and without constipation (G4, n = 23). In addition to characterizing subjects' clinical demographics, medication intake, disease severity levels, salivary cytokine levels [TNF-α, interleukin (IL)-1ß, IL-6, IL-8, IL-10], and Caregiver Priorities and Child Health Index of Life with Disabilities (CPCHILD). Statistical significance was evaluated by Shapiro-Wilks, Student's T-Test, ANOVA, and ANCOVA analysis. Results: Salivary proinflammatory cytokines were highly correlated with the severe form of gut constipation in G1 (P < 0.001), and out of all cytokines IL-1ß levels demonstrated highest correlation with all gut constipation (P < 0.05). A significant relationship was found between the type of medication, in which subjects taking Gamma-Aminobutyric Acid (GABA) and GABA+ (GABA in association with other medication) were more likely to be constipated than the other groups (P < 0.01). Cleary salivary inflammatory levels and gut constipation were correlated, and impacted QOL of CP subjects. G1 presented a lower QOL mean score of CPCHILD (49.0 ± 13.1) compared to G2 (71.5 ± 16.7), when compared to G3 (88.9 ± 7.5), and G4 (95.5 ± 5.0) (P < 0.01). We accounted for gingival bleeding as a cofounder of oral inflammation, and here were no differences among groups regarding gender (P = 0.332) and age (P = 0.292). Conclusions: Collectively, the results suggest that saliva inflammatory levels were linked to gut constipation, and that the clinical impact of medications that controlled gut was reliably monitored via oral cytokine levels, providing reliable and non-invasive information in precision diagnostics.

Does the neuromotor abnormality type affect the salivary parameters in individuals with cerebral palsy?

BACKGROUND: Previous studies reported alterations in salivary flow rate and biochemical parameters of saliva in cerebral palsy (CP) individuals; however, none of these considered the type of neuromotor abnormality among CP individuals, thus it remains unclear whether the different anatomical and extended regions of the brain lesions responsible for the neurological damage in CP might include disruption of the regulatory mechanism of saliva secretion as part of the encephalopathy. The aim of this study was to evaluate salivary flow rate, pH and buffer capacity in saliva of individuals with CP, aged 3-16 years, with spastic neuromotor abnormality type and clinical patterns of involvement. METHODS: Sixty-seven individuals with CP spasticity movement disorder, were divided in two groups according to age (3-8- and 9-16-years-old) and compared with 35 sibling volunteers with no neurological damage, divided in two groups according to age (3-8- and 9-16-years-old). Whole saliva was collected under slight suction and pH and buffer capacity were determined using a digital pH meter. Buffer capacity was measured by titration using 0.01N HCL, and flow rate was calculated in ml/min. RESULTS: In both age groups studied, whole saliva flow rate, pH and buffer capacity were significantly lower in the spastic CP group (P<0.05). The clinical patterns of involvement did not influence the studied parameters. CONCLUSION: These findings show that individuals with spastic cerebral palsy present lower salivary flow rate, pH and buffer capacity that can increase the risk of oral disease in this population.

Salivary lead concentration in dental caries among normal and children with cerebral palsy.

INTRODUCTION: Cerebral palsy (CP) is a neurological disability the exact cause of which is not known. Exposure to toxic elements, environmental pollutants, and various teratogens such as lead, either prenatal or postnatal, can be a risk factor for this neurologic disability. CP children have poor neuromuscular coordination, exposing them to increased risk of oral diseases such as drooling of saliva, periodontal diseases, dental caries, and malocclusion. There are less studies comparing lead concentration in CP children, as it can be one of the contributing factors to dental caries. AIMS AND OBJECTIVES: The present study was to estimate and compare the salivary lead level in normal and neurologically disabled children and to correlate salivary lead level with dental caries in both normal and neurologically disabled children. MATERIALS AND METHODS: A study on 45 children reporting to CP rehabilitation center and 41 normal children taken from the Outpatient Department of the Pediatric and Preventive Dentistry. P.M.N.M. Dental College and Hospital, Bagalkot, Karnataka. All children were between the age group of 5-12 years. In all individuals, a thorough medical history and dental examination were done. The age, state of dentition, and the level of caries in all individuals were determined by the same examiner, and salivary lead concentrations were determined using atomic absorption spectrophotometer. All children were divided into four groups depending on the presence or absence of dental caries. Group 1 consisted of 20 normal children with dental caries, Group 2 consisted of 21 normal children without dental caries, Group 3 consisted of 25 CP children with dental caries, and Group 4 consisted of 20 CP children without dental caries. RESULTS: Were recorded tabulated and statistically analyzed. CP children had high decayed, missing, and filled teeth/def scores, dental caries, and salivary lead concentration as compared to normal children. Statistically significant correlation was obtained between the dental caries and saliva lead concentration. The lead concentration was directly proportional to the carious status. CONCLUSION: CP children are more prone to dental caries and increased salivary lead concentration which could be a cause as well as the effect of CP.

Comparison of electrolyte concentrations in whole saliva of individuals with and without cerebral palsy.

OBJECTIVE: To analyze sodium, potassium, calcium, phosphorus, and magnesium concentrations in whole saliva of adolescents with cerebral palsy (CP) and compare them with those of normal individuals. METHOD AND MATERIALS: Thirty-six adolescents with CP aged 12.0 +/- 1.8 years were compared with 36 healthy age-matched individuals. Saliva was collected under slight suction. Electrolyte concentrations were determined by inductively coupled argon plasma with atomic emission spectrometry. RESULTS: Significant differences were observed with matched and unmatched flow rates. Sodium concentrations were significantly decreased, whereas potassium levels were significantly increased in the CP group. No statistically significant differences were observed regarding phosphorus, magnesium, and calcium concentrations between adolescents with CP and controls. CONCLUSION: Our results suggest that there is an alteration in the sodium and potassium concentration in whole saliva of individuals with CP.

Microstructure and chemical composition of primary teeth in children with Down syndrome and cerebral palsy.

This study was designed to test the hypothesis that prenatal growth insults leave permanent signs in the developing primary teeth that can be identified in later life. To test this hypothesis we examined exfoliated and extracted lower second primary molars of children with Down syndrome (DS) and cerebral palsy (CP). Teeth of children with no adverse medical history were used as a control group. Informed consent of parents and children was obtained in all cases. On each tooth two thin sections were cut, one bisecting the mesial cusps and one bisecting the distal cusps. Using a light microscope, the width of prenatal enamel and postnatal enamel was measured on each section at standardized locations from the dentin-enamel junction (DEJ) with the neonatal line used to distinguish between prenatal and postnatal enamel. Chemical analysis of each section was carried out using an energy dispersive spectrophotometer (ESR). The Ca/P ratios of enamel and dentin for each cusp were calculated and intercusp and intergroup differences analysed using non-parametric statistical tests. The results showed that significantly less enamel was laid down prenatally in DS and CP teeth than in the control group and that the enamel of the mesial cusps in these groups was less highly mineralised than that of the controls. The results also showed that in DS teeth growth and mineralisation of all cusps was affected. Based on these findings we propose that analysis of exfoliated deciduous teeth in developmentally challenged children may help in identifying the onset and severity of growth insults in utero and its impact on later development.

Salivary osmolality in individuals with cerebral palsy.

OBJECTIVE: To measure the salivary flow rate, osmolality, electrolyte and total protein concentrations in individuals with cerebral palsy (CP). DESIGN: Thirty-eight individuals with CP were divided according to the neuromotor abnormality type (total, spastic and dyskinectic) and compared to 22 nondisabled children (control group). Whole saliva was collected under slight suction. The salivary parameters studied were salivary flow rate, osmolality, sodium, potassium, chloride and total protein concentrations. RESULTS: CP individuals, with both neuromotor abnormality types (spastic and dyskinectic), presented an increase in salivary osmolality, total protein, potassium and chloride concentrations compared to the control group (p<0.05). Moreover, a reduction in salivary flow rate was verified in spastic individuals (p<0.05). CONCLUSION: The reduction in salivary flow rate and increase in osmolality, total protein and electrolyte concentrations of saliva from cerebral palsy individuals could be caused by hypohydration status.