Multidimensional analysis of human intestinal fluid composition.

The oral administration of solid dosage forms is the commonest method to achieve systemic therapy and relies on the drug's solubility in human intestinal fluid (HIF), a key factor that influences bioavailability and biopharmaceutical classification. However, HIF is difficult to obtain and is known to be variable, which has led to the development of a range of simulated intestinal fluid (SIF) systems to determine drug solubility in vitro. In this study we have applied a novel multidimensional approach to analyse and characterise HIF composition using a published data set in both fasted and fed states with a view to refining the existing SIF approaches. The data set provided 152 and 172 measurements of five variables (total bile salt, phospholipid, total free fatty acid, cholesterol and pH) in time-dependent HIF samples from 20 volunteers in the fasted and fed state, respectively. The variable data sets for both fasted state and fed state are complex, do not follow normal distributions but the amphiphilic variable concentrations are correlated. When plotted 2-dimensionally a generally ellipsoid shaped data cloud with a positive slope is revealed with boundaries that enclose published fasted or fed HIF compositions. The data cloud also encloses the majority of fasted state and fed state SIF recipes and illustrates that the structured nature of design of experiment (DoE) approaches does not optimally cover the variable space and may examine media compositions that are not biorelevant. A principal component analysis in either fasted or fed state in combination with fitting an ellipsoid shape to enclose the data results in 8 points that capture over 95% of the compositional variability of HIF. The variable's average rate of concentration change in both fasted state and fed state over a short time scale (10 min) is zero and a Euclidean analysis highlights differences between the fasted and fed states and among individual volunteers. The results indicate that a 9-point DoE (8 + 1 central point) could be applied to investigate drug solubility in vitro and provide statistical solubility limits. In addition, a single point could provide a worst-case solubility measurement to define the lowest biopharmaceutical classification boundary or for use during drug development. This study has provided a novel description of HIF composition. The approach could be expanded in multiple ways by incorporation of further data sets to improve the statistical coverage or to cover specific patient groups (e.g., paediatric). Further development might also be possible to analyse information on the time dependent behaviour of HIF and to guide HIF sampling and analysis protocols.

Intratracheal LPS administration attenuates the acute hypoxic ventilatory response: Role of brainstem IL-1ß receptors.

Perinatal inflammation and infection are commonly associated with various respiratory morbidities in preterm infants including apnea of prematurity. In this study, we investigated whether pulmonary inflammation via intra-tracheal micro-injection of lipopolysaccharide (LPS) into neonatal rats modifies respiratory neural control via an IL-1ß receptor-dependent mechanism. Prior to an intra-tracheal micro-injection of LPS (1mg/kg), 10day old (Postnatal age, P10) rats received an intraperitoneal (i.p.) or intracisternal (i.c.) micro-injection of the IL-1ß receptor antagonist AF12198. Whole-body plethysmography was performed two hours later to assess the magnitude of the acute hypoxic (HVR) and hypercapnic (HCVR) ventilatory responses. Intra-tracheal LPS dose-dependently attenuated the acute HVR compared to saline (control) treated rats, whereas the HCVR was not affected. Pre-treatment with an i.c. (but not i.p.) micro-injection of AF12198 15min prior to LPS prevented the attenuated HVR. These data indicate that intrapulmonary inflammation affects brainstem respiratory neural pathways mediating the ventilatory response to acute hypoxia via an IL-1ß-dependent pathway. These findings are relevant to our understanding of the way that pulmonary inflammation may affect central neural mechanisms of respiratory insufficiency commonly seen in preterm infants.

Changes in carotid body and nTS neuronal excitability following neonatal sustained and chronic intermittent hypoxia exposure.

We investigated whether pre-treatment with neonatal sustained hypoxia (SH) prior to chronic intermittent hypoxia (SH+CIH) would modify in vitro carotid body (CB) chemoreceptor activity and the excitability of neurons in the caudal nucleus of the solitary tract (nTS). Sustained hypoxia followed by CIH exposure simulates an oxygen paradigm experienced by extremely premature infants who developed persistent apnea. Rat pups were treated with 5 days of SH (11% O2) from postnatal age 1 (P1) followed by 10 days of subsequent chronic intermittent hypoxia (CIH, 5% O2/5 min, 8 h/day, between P6 and P15) as described previously (Mayer et al., Respir. Physiol. Neurobiol. 187(2): 167-75, 2013). At the end of SH+CIH exposure (P16), basal firing frequency was enhanced, and the hypoxic sensory response of single unit CB chemoafferents was attenuated. Further, basal firing frequency and the amplitude of evoked excitatory post-synaptic currents (ESPC's) of nTS neurons was augmented compared to age-matched rats raised in normoxia. These effects were unique to SH+CIH exposure as neither SH or CIH alone elicited any comparable effect on chemoafferent activity or nTS function. These data indicated that pre-treatment with neonatal SH prior to CIH exposure uniquely modified mechanisms of peripheral (CB) and central (nTS) neural function in a way that would be expected to disturb the ventilatory response to acute hypoxia.

Erythropoietin attenuates loss of potassium chloride co-transporters following prenatal brain injury.

Therapeutic agents that restore the inhibitory actions of γ-amino butyric acid (GABA) by modulating intracellular chloride concentrations will provide novel avenues to treat stroke, chronic pain, epilepsy, autism, and neurodegenerative and cognitive disorders. During development, upregulation of the potassium-chloride co-transporter KCC2, and the resultant switch from excitatory to inhibitory responses to GABA guide the formation of essential inhibitory circuits. Importantly, maturation of inhibitory mechanisms is also central to the development of excitatory circuits and proper balance between excitatory and inhibitory networks in the developing brain. Loss of KCC2 expression occurs in postmortem samples from human preterm infant brains with white matter lesions. Here we show that late gestation brain injury in a rat model of extreme prematurity impairs the developmental upregulation of potassium chloride co-transporters during a critical postnatal period of circuit maturation in CA3 hippocampus by inducing a sustained loss of oligomeric KCC2 via a calpain-dependent mechanism. Further, administration of erythropoietin (EPO) in a clinically relevant postnatal dosing regimen following the prenatal injury protects the developing brain by reducing calpain activity, restoring oligomeric KCC2 expression and attenuating KCC2 fragmentation, thus providing the first report of a safe therapy to address deficits in KCC2 expression. Together, these data indicate it is possible to reverse abnormalities in KCC2 expression during the postnatal period, and potentially reverse deficits in inhibitory circuit formation central to cognitive impairment and epileptogenesis.

Advanced BMP gene therapies for temporal and spatial control of bone regeneration.

Spatial and temporal patterns of bone morphogenetic protein (BMP) signaling are crucial to the assembly of appropriately positioned and shaped bones of the face and head. This review advances the hypothesis that reconstitution of such patterns with cutting-edge gene therapies will transform the clinical management of craniofacial bone defects attributed to trauma, disease, or surgical resection. Gradients in BMP signaling within developing limbs and orofacial primordia regulate proliferation and differentiation of mesenchymal progenitors. Similarly, vascular and mesenchymal cells express BMPs in various places and at various times during normal fracture healing. In non-healing fractures of long bones, BMP signaling is severely attenuated. Devices that release recombinant BMPs promote healing of bone in spinal fusions and, in some cases, of open fractures, but cannot control the timing and localization of BMP release. Gene therapies with regulated expression systems may provide substantial improvements in efficacy and safety compared with protein-based therapies. Synthetic gene switches, activated by pharmacologics or light or hyperthermic stimuli, provide several avenues for the non-invasive regulation of the expression of BMP transgenes in both time and space. Through new gene therapy platforms such as these, active control over BMP signaling can be achieved to accelerate bone regeneration.

Physiologic basis for intermittent hypoxic episodes in preterm infants.

Intermittent hypoxic episodes are typically a consequence of immature respiratory control and remain a troublesome challenge for the neonatologist. Furthermore, their frequency and magnitude are commonly underestimated by clinically employed pulse oximeter settings. In extremely low birth weight infants the incidence of intermittent hypoxia [IH] progressively increases over the first 4 weeks of postnatal life, with a subsequent plateau followed by a slow decline beginning at weeks six to eight. Over this period of unstable respiratory control, increased oxygen-sensitive peripheral chemoreceptor activity has been associated with a higher incidence of apnea of prematurity. In contrast, infants with bronchopulmonary dysplasia [chronic neonatal lung disease] exhibit decreased peripheral chemosensitivity, although the effect on respiratory stability in this population is unclear. Such episodic hypoxia/reoxygenation in early life has the potential to sustain a proinflammatory cascade with resultant multisystem, including respiratory, morbidity. Therapeutic approaches for intermittent hypoxic episodes comprise careful titration of baseline or supplemental inspired oxygen as well as xanthine therapy to prevent apnea of prematurity. Characterization of the pathophysiologic basis for such intermittent hypoxic episodes and their consequences during early life is necessary to provide an evidence-based approach to their management.

Modulation of gastric pH by a buffered soluble effervescent formulation: A possible means of improving gastric tolerability of alendronate.

Gastrointestinal side-effects of alendronate (ALN) are believed to be associated with oesophageal lodging of tablets and perhaps reflux of gastric contents with alendronate under strongly acidic pH conditions. This leads to unfavourable posture restrictions when dosing. This clinical study evaluated gastric emptying and gastric pH after administration of Fosamax(®) tablets and a novel effervescent ALN formulation with a high buffering capacity. This novel formulation, EX101, was developed to potentially improve gastric tolerance. Gastric pH was monitored by nasogastric probes. Gastric emptying was determined simultaneously by scintigraphic imaging of (99m)Tc-DTPA labelled formulations. Both formulations tested rapidly cleared the oesophagus and there were no statistically significant or physiologically relevant differences in gastric emptying times. Mean pH at time to 50% gastric emptying of the radiolabel was significantly higher in EX101-treated subjects compared to those treated with Fosamax(®). At time to 90% gastric emptying of the radiolabel, mean pH values were comparable. Mucosal exposure to ALN at pH less than 3 is irritating to gastro-oesophageal tissue. Ingestion of Fosamax(®) resulted in ALN being present in the stomach at a pH below 3 within minutes. EX101 minimised the possibility of exposing the oesophagus (in case of reflux) to acidified ALN.

Comparison of a novel fast-dissolving acetaminophen tablet formulation (FD-APAP) and standard acetaminophen tablets using gamma scintigraphy and pharmacokinetic studies.

CONTEXT: Acetaminophen (paracetamol, APAP) is widely used to relieve mild-to-moderate pain and reduce fever. Absorption of the drug can be impacted by dosage form; this may have implications for pain relief in some individuals, potentially accounting for suboptimal efficacy in analgesia. OBJECTIVE: To assess the disintegration and dissolution of a new fast-dissolving acetaminophen tablet formulation (FD-APAP) and the impact on pharmacokinetic and pharmacodynamic parameters. MATERIALS AND METHODS: Two randomized, single-center, open-label, single-dose, two-way crossover studies in healthy subjects to compare FD-APAP (2 × 500 mg tablets) with standard acetaminophen (2 × 500 mg tablets). Gamma scintigraphy was used to assess tablet disintegration (Study 1, N = 24), and plasma profiles were evaluated in the fasted state (Study 2, N = 40). RESULTS: In Study 1, the mean time to complete disintegration (12.9 vs. 69.6 min, P < 0.0001) and onset of disintegration were both significantly faster with FD-APAP than with standard acetaminophen (P < 0.0001). For Study 2, median T(max) was significantly faster for FD-APAP (0.50 vs. 0.67 h, P < 0.01) and AUC(0-30 min) was significantly greater (4.51 vs. 2.74, P < 0.05). AUC(0-t) and AUC(0-inf) were comparable between the two study treatments. DISCUSSION: Despite the absence of comparative clinical data, the FD-APAP formulation may be expected to overcome some of the issues associated with the slow and variable absorption of standard acetaminophen tablet formulations, improving therapeutic outcome and avoiding the need to switch to alternative therapeutic options. CONCLUSION: Compared with standard acetaminophen, the FD-APAP formulation results in significantly faster onset of disintegration and more rapid absorption.

Small-molecule inhibitors of IL-2/IL-2R: lessons learned and applied.

The IL-2:IL-2R protein-protein interaction is of central importance to both healthy and diseased immune responses, and is one of the earliest examples of successful small-molecule inhibitor discovery against this target class. Drug-like inhibitors of IL-2 have been identified through a combination of fragment discovery, structure-based design, and medicinal chemistry; this discovery approach illustrates the importance of using a diverse range of complementary screening methods and analytical tools to achieve a comprehensive understanding of molecular recognition. The IL-2 story also provides insight into the dynamic nature of protein-protein interaction surfaces, their potential druggability, and the physical and chemical properties of effective small-molecule ligands. These lessons, from IL-2 and similar discovery programs, underscore an increasing awareness of the principles governing the development of drugs for protein-protein interactions.

Source-pathway separation of multiple contaminants during a rainfall-runoff event in an artificially drained agricultural watershed.

A watershed's water quality is influenced by contaminant-transport pathways unique to each landscape. Accurate information on contaminant-pathways could provide a basis for mitigation through well-targeted approaches. This study determined dynamics of nitrate-N, total P, Escherichia coli, and sediment during a runoff event in Tipton Creek, Iowa. The watershed, under crop and livestock production, has extensive tile drainage discharging through an alluvial valley. A September 2006 storm yielded 5.9 mm of discharge during the ensuing 7 d, which was monitored at the outlet (19,850 ha), two tile-drainage outfalls (total 1856 ha), and a runoff flume (11 ha) within the sloped valley. Hydrograph separations indicated 13% of tile discharge was from surface intakes. Tile and outlet nitrate-N loads were similar, verifying subsurface tiles dominate nitrate delivery. On a unit-area basis, tile total P and E. coli loads, respectively, were about half and 30% of the outlet's; their rapid, synchronous timing showed surface intakes are an important pathway for both contaminants. Flume results indicated field runoff was a significant source of total P and E. coli loads, but not the dominant one. At the outlet, sediment, P, and E. coli were reasonably synchronous. Radionuclide activities of (7)Be and (210)Pb in suspended sediments showed sheet-and-rill erosion sourced only 22% of sediment contributions; therefore, channel sources dominated and were an important source of P and E. coli. The contaminants followed unique pathways, necessitating separate mitigation strategies. To comprehensively address water quality, erosion-control and nitrogen-management practices currently encouraged could be complemented by buffering surface intakes and stabilizing stream banks.

In situ measurement of spectral changes in the anterior eye following application of ultraviolet-absorbing compounds.

The ocular structures are very sensitive to damage from ultraviolet (UV) radiation, exposure is linked to corneal and conjunctival damage, cataract formation and may also be implicated in the aetiology of age-related macular degeneration. These structures are usually protected by wearing suitable eyeglasses and goggles. An alternative to conventional eyeglasses/goggles is the concept of "liquid sunglasses" which involve the topical application of eye drops that are designed to block harmful UV radiation reaching the sensitive ocular surfaces. The evaluation of such compounds directly applied to the eye surface requires in situ measurements to compare the efficacy of different formulations. A novel ocular spectrometer system has been used to evaluate changes in the transmission of ultraviolet (UV) radiation through the anterior eye following topical application of candidate UV-absorbing formulations. The key feature of the system is the ability to propagate a beam of light tangentially through the anterior eye using a compact, hand-held lens assembly incorporating UV-transmitting optical fibres. A range of formulations containing UV-absorbing compounds were topically applied to ex vivo rabbit eyes. Significant increases in the absorption of the UV spectrum were detected in seven of the eight formulations studied, demonstrating the potential of this measurement technique in the evaluation of formulations developed as potential topical ocular sunscreens.

Exploring microdischarges for portable sensing applications.

This paper describes the use of microdischarges as transducing elements in sensors and detectors. Chemical and physical sensing of gases, chemical sensing of liquids, and radiation detection are described. These applications are explored from the perspective of their use in portable microsystems, with emphasis on compactness, power consumption, the ability to operate at or near atmospheric pressure (to reduce pumping challenges), and the ability to operate in an air ambient (to reduce the need for reservoirs of carrier gases). Manufacturing methods and performance results are described for selected examples.

Do gastrointestinal transit parameters influence the pharmacokinetics of gefitinib?

The selective EGFR tyrosine kinase inhibitor, gefitinib has been shown to be active against certain human carcinomas. It had been noted that a proportion of volunteers consistently had lower gefitinib exposure following oral administration. The shape of the elimination profile in this subset was also different, showing a monophasic elimination pattern rather than the biphasic pattern observed in the majority of subjects. A gamma scintigraphic study was conducted to examine the relationship of gastrointestinal transit and drug absorption in a cohort of rapid clearance subjects (n=5) and normal profile volunteers (n=7). The fasted volunteer panel received a 250 mg gefitinib tablet labelled with [(111)In]-DTPA together with 240 mL [(99m)Tc]-labelled water. The rapid clearance cohorts were shown to have a faster mean gastric emptying T90 (37 min vs 74 min) and shorter small intestinal transit time (156 min vs 204 min), resulting in an earlier colonic arrival time (181 min vs 244 min). Mean plasma C(max) was lower (99.2 ng/mL vs 116 ng/mL) and AUC almost half in the rapid clearance group (2162+/-81 ngh/mL vs 4996+/-64 ngh/mL). These data suggest that gastrointestinal transit parameters play a role in the differences in the rapid clearance profile group, also contributing to the biphasic to monophasic switch. However, historical data show, at the recommended dose of 250 mg/day steady-state plasma concentrations adequate for clinical benefit are achieved in patients with non-small cell lung cancer.

Staphylococcus aureus colonization of anterior nares of school going children.

OBJECTIVE: To determine the colonization rates of S. aureus in anterior nares of school going children, evaluate the antimicrobial resistance of such isolates against various antibiotics. METHODS: Nasal swabs taken for S. aureus in 392 healthy school going children aged between 5 and 15 yr belonging to three schools surrounding Kamineni Institute of Medical Sciences, Narketpally, Andhra Pradesh, India. Swabs were inoculated in to Mannitol Salt Agar (MSA) and incubation at 37 degrees C for 24hr, Staphylococcus aureus isolates were identified by standard microbiological methods such as Gram's stain, catalase and coagulase. MICs were determined by Agar dilution technique against Vancomycin, Ciprofloxacin, Sparfloxacin, Sparfloxacin beta Cyclodextrin. Antimicrobial resistance patterns of all the isolates against Oxacillin (1 microg) Penicillin (10 units), Ampicillin (10 microg), Co-trimoxazole (23.75 microg), Erythromycin (15 microg), Tetracycline (30 microg) and Gentamicin (10 microg) were tested using Kirby-Baeur disk diffusion method in accordance with CLSI standards. RESULTS: Of the 392 samples screened 63(16%) showed the growth of Staphylococcus aureus. 12(19%) isolates were found to be MRSA. Antimicrobial susceptibility testing using Agar dilution method against Vancomycin, ciprofloxacin, Sparfloxacin, and Sparfloxacin beta Cyclodextrin revealed MICs in the range of 0.52 microg/ml, 0.51 microg/ml, 0.5 microg/ml and <0.0312-0.250 microg/ml respectively. Disk diffusion method showed that all the isolates were resistant to Penicillin and Ampicillin. A resistance of 14.3%, 25% and 22.2% was observed against Co-trimoxazole, Erythromycin and Tetracycline respectively. Gentamiicin was the only antibiotic against which most of the isolates were sensitive. CONCLUSION: Our results suggests that healthy school going children under 16 yr of age are potential carriers of S. aureus and in particular MRSA and multi-drug resistant strains.

Composition-function relationships during IL-1-induced cartilage degradation and recovery.

OBJECTIVE: To examine the relationships between biochemical composition and mechanical properties of articular cartilage explants during interleukin-1 (IL-1)-induced degradation and post-exposure recovery. DESIGN: Bovine articular cartilage explants were cultured for up to 32 days with or without 20 ng/mL IL-1. The dynamic shear modulus |G*(dyn)| and equilibrium and dynamic unconfined compression moduli (E(equil) and |E*(dyn)|) were measured at intervals throughout the culture period. In a subsequent recovery study, explants were cultured for 4 days with or without 20ng/mL IL-1 and for an additional 16 days in control media. The dynamic moduli |E*(dyn)| and |G*(dyn)| were measured at intervals during degeneration and recovery. Conditioned media and explant digests were assayed for sulfated glycosaminoglycans (sGAG) and collagen content. RESULTS: Continuous IL-1 stimulation triggered progressive decreases in E(equil), |E*(dyn)|, and |G*(dyn)| concomitant with the sequential release of sGAG and collagen from the explants. Brief IL-1 exposure resulted in a short release of sGAG but not collagen, followed by a gradual and incomplete repopulation of sGAG. The temporary sGAG depletion was associated with decreases in both |E*(dyn)| and |G*(dyn)| which also recovered after removal of IL-1. During IL-1-induced degradation and post-exposure recovery, explant mechanical properties correlated well with tissue sGAG concentration. CONCLUSIONS: As previously shown for developing cartilages and engineered cartilage constructs, cytokine-induced changes in sGAG concentration (i.e., fixed charge density) are coincident with changes in compressive and shear properties of articular cartilage. Further, recovery of cartilage mechanical properties can be achieved by relief from proinflammatory stimuli and subsequent restoration of tissue sGAG concentration.

Scintigraphic evaluation of colon targeting pectin-HPMC tablets in healthy volunteers.

The in vivo evaluation of colon-targeting tablets was conducted in six healthy male volunteers. A pectin-hydroxypropyl methylcellulose coating was compressed onto core tablets labelled with 4MBq (99m)Tc-DTPA. The tablets released in the colon in all subjects; three in the ascending colon (AC) and three in the transverse colon (TC). Tablets that released in the TC had reached the AC before or just after food (Group A). The other three tablets released immediately upon AC entry at least 1.5h post-meal (Group B). Release onset for Group B was earlier than Group A (343min vs 448min). Group B tablets exhibited a clear residence period at the ileocaecal junction (ICJ) which was not observed in Group A. Prolonged residence at the ICJ is assumed to have increased hydration of the hydrogel layer surrounding the core tablet. Forces applied as the tablets progressed through the ICJ may have disrupted the hydrogel layer sufficiently to initiate radiolabel release. Conversely, Group A tablets moved rapidly through the AC to the TC, possibly minimising contact times with water pockets. Inadequate prior hydration of the hydrogel layer preventing access of pectinolytic enzymes and reduced fluid availability in the TC may have retarded tablet disintegration and radiolabel diffusion.

Articular chondrocytes derived from distinct tissue zones differentially respond to in vitro oscillatory tensile loading.

OBJECTIVE: The cell morphology, gene expression, and matrix synthesis of articular chondrocytes are known to vary with depth from the tissue surface. The objective of this study was to investigate if chondrocytes from different zones respond to in vitro oscillatory tensile loading in distinct ways and whether tensile strain, which is most prevalent near the articular surface, would preferentially stimulate superficial zone chondrocytes. DESIGN: Chondrocytes were separately isolated from the superficial, middle, and deep zones of articular cartilage and seeded into three-dimensional fibrin hydrogel constructs. An intermittent protocol of oscillatory tensile loading was applied for 3 days, and the effects on extracellular matrix (ECM) synthesis were assessed by measuring the incorporation of radiolabed precursors, size exclusion gel chromatography, and western blotting. RESULTS: Tensile loading was found to be a potent stimulus for proteoglycan synthesis only in superficial zone chondrocytes. Although overall biosynthesis rates by deep zone chondrocytes were unaffected by tensile loading, the molecular characteristics of proteins and proteoglycans released to the culture medium were significantly altered so as to resemble those of superficial zone chondrocytes. CONCLUSIONS: Oscillatory tensile loading differentially affected subpopulations of articular chondrocytes in three-dimensional fibrin hydrogel constructs. Cells isolated from deeper regions of the tissue developed some characteristics of superficial zone chondrocytes after exposure to tensile loading, which may indicate an adaptive response to the new mechanical environment. Understanding how exogenous mechanical stimuli can differentially influence chondrocytes from distinct tissue zones will yield important insights into mechanobiological processes involved in cartilage tissue development, maintenance, disease, and repair.

Characterization of proteoglycan production and processing by chondrocytes and BMSCs in tissue engineered constructs.

OBJECTIVE: The goal of this study was to characterize the proteoglycan (PG) production and processing by bone marrow stromal cells (BMSCs) within a tissue engineered construct. METHODS: Bovine BMSCs and articular chondrocytes (ACs) were isolated from an immature calf, seeded into agarose gels, and cultured up to 32 days in the presence of TGF-beta1. The localization of various PGs was examined by immunofluorescence and histological staining. The role of proteolytic enzymes in construct development was further investigated by examining the effects of aggrecanase and MMP inhibitors on PG accumulation, aggrecan processing, and construct mechanics. RESULTS: BMSCs developed a matrix rich in sulfated-glycosaminoglycans (sGAG) and full-length aggrecan, but had low levels of versican. The BMSC constructs had less collagen II and aggrecan compared to the AC constructs cultured under identical conditions. AC constructs also had high levels of pericellular collagen VI, while BMSCs had a pericellular matrix containing little collagen VI and greater levels of decorin, biglycan, and fibronectin. Treatment with the aggrecanase inhibitor did not affect sGAG accumulation or the dynamic moduli of the BMSC constructs. The MMP inhibitor slightly but significantly inhibited sGAG accumulation and lowered the dynamic moduli of BMSC constructs. CONCLUSIONS: The results of this preliminary study indicate that long-term culture of BMSCs with TGF-beta1 promotes the development of an aggrecan-rich matrix characteristic of native articular cartilage; however, BMSCs accumulate significantly lower levels of sGAG and assemble distinct pericellular microenvironments compared to ACs. PG turnover does not appear to play a major role in the development of tissue engineered cartilage constructs by BMSCs.

Development and in vitro/in vivo evaluations of bioadhesive buccal tablets for nicotine replacement therapy.

Buccal bioadhesive tablet formulations of nicotine hydrogen tartrate (NHT) for nicotine replacement therapy (NRT) were developed using chitosan and carbomer at different ratios. Magnesium hydroxide was incorporated into the formulations as pH increasing agent. In vitro release and bioadhesion properties of the tablets were investigated. Release of NHT from the tablets was increased with the increasing amount of chitosan in formulations whilst the bioadhesion of the tablet was decreased. In vivo studies were carried out in healthy, non-smoker volunteers in comparison to a commercially available transdermal patch. Plasma nicotine and cotinine levels were determined using gas chromatography-mass spectrophotometry. No significant difference was found between the maximum plasma nicotine concentrations (Cmax) obtained with the buccal tablet and the transdermal patch (p > 0.05). Time to reach the Cmax was 2.9 +/- 0.2 h and 11.5 +/- 1.3 h, and AUC0-24 values were 59.3 +/- 5.1 ng x h x mL(-1) (0-12 h) and 204.1 +/- 31.2 ng x h x mL(-1) for buccal tablet and transdermal patch, respectively.