Bioengineered tooth emulation systems for regenerative and pharmacological purposes.

Genetic conditions, traumatic injuries, carious lesions and periodontal diseases are all responsible for dental pathologies. The current clinical approaches are based on the substitution of damaged dental tissues with inert materials, which, however, do not ensure full physiological recovery of the teeth. Different populations of dental mesenchymal stem cells have been isolated from dental tissues and several attempts have already been made at using these stem cells for the regeneration of human dental tissues. Despite encouraging progresses, dental regenerative therapies are very far from any clinical applications. This is tightly connected with the absence of proper platforms that would model and faithfully mimic human dental tissues in their complexity. Therefore, in the last decades, many efforts have been dedicated for the development of innovative systems capable of emulating human tooth physiology in vitro. This review focuses on the use of in vitro culture systems, such as bioreactors and "organ-on-a-chip" microfluidic devices, for the modelling of human dental tissues and their potential use for dental regeneration and drug testing.

A bio-degradable synthetic membrane to treat superficial and deep second degree burn wounds in adults and children - 4 year experience.

INTRODUCTION: A new bio-degradable synthetic membrane was recently introduced to treat second degree burns in adults and pediatric patients. OBJECTIVE: To assess complications and outcomes using this absorbable synthetic membrane to treat second degree burns. METHODS: 229 burn patients, 138 pediatric, with superficial and deep second -degree wounds, treated with the absorbable synthetic membrane (Suprathel®, Polymedics, Denkendorf, Germany) were included in this study. Patients were treated under anesthesia or moderate sedation. The wound bed was prepared by using either rough debridement or dermabrasion excision. After hemostasis, the membrane was applied to the wound with an outer layer dressing of fatty gauze, bridal veil, absorptive gauze and an ACE® wrap. The outer dressing was removed every one to four days, depending on exudate, in order to closely follow the wound through the translucent membrane and fatty gauze layers. After complete epithelialization, the dressing separated and could be removed. The study focused on the need for subsequent grafting, healing time, patient pain level, hypertrophic scarring and rate of infection. RESULTS: All wounds in this study that were treated with Suprathel® healed without grafting. The average TBSA (Total Body Surface Area) was 8.9% (1%-60%). Average time to healing was 13.7 days for ≥ 90% epithelialization with 11.9 days for pediatric patients versus 14.7 days for adults. Throughout the treatment period, the average pain level was 1.9 on a 10-point scale. 27 patients developed hypertrophic scarring in some areas (11.7%). Average Length of stay (LOS) was 6.9 days. The rate of infection was 3.8% (8/229). Failure or progression to full thickness in part of the wounds was 5.2% (12/229). CONCLUSION: In treating second degree burn wounds, this membrane provides a simple, effective solution alternative with good outcomes and less pain than conventional and previously studied treatment options in the same institution. Fewer dressing changes and easier overall management of the wounds contribute to its favorable profile.

The effects of ageing on dental pulp stem cells, the tooth longevity elixir.

Stem cells are essential for tissue homeostasis and regeneration throughout the lifespan of multicellular organisms. The decline in stem cell function during advanced age is associated with a reduced regenerative potential of tissues that leads to an increased frequency of diseases. Age-related changes also occur in the dental pulp that represents a reliable model tissue, with high regenerative capability, for studying senescence mechanisms. However, little information is available concerning the effects of ageing on dental stem-cell function. In this mini-review, recent data on how the molecular and functional alterations that accumulate in stem cell populations during ageing result in modifications of dental pulp physiology are discussed. Changes that accumulate during ageing such as how reduction of pulp chamber volume, decreased vascular supply and modifications to the stem cell niches affect stem cell functions and, therefore, dental pulp regenerative potential in response to various stressful agents. Dental pulp cells from aged individuals are still metabolically active and secrete pro-inflammatory and matrix-degrading molecules. Furthermore, miRNAs and exosomes derived from dental pulp stem cells constitute an attractive source of nanovesicles for the treatment of age-related dental pathologies. Further investigation of the epigenetic alterations in dental pulp stem cells, accumulating during ageing, might reveal crucial information for potential stem cell-based therapeutic approaches in the elderly.

Investigation of orofacial stem cell niches and their innervation through microfluidic devices.

Stem cell-based mediated therapies represent very promising approaches for tissue regeneration and are already applied with success in clinics. These therapeutic approaches consist of the in vitro manipulation of stem cells and their consequent administration to patients as living and dynamic biological agents. Nevertheless, the deregulation of stem cells function might result in the generation of pathologies such as tumours or accelerated senescence. Moreover, different stem cells sources are needed for regeneration of specific tissues. It is thus fundamental to understand the mechanisms regulating the physiology of stem cells. Microfluidic technology can be used to mimic in vivo scenarios and allow the study of stem cell physiology at both single cell and whole stem cell niche levels.This review focuses on the potential sources of stem and progenitor cells for orofacial regeneration and the use of microfluidic technologies for the study of stem cells behaviour and stem cell niches, in the light of regenerative medicine.

Effects of MF-268, a new cholinesterase inhibitor, on acetylcholine and biogenic amines in rat cortex.

MF-268 bitartrate [(3a S, 8a R)-1,2,3,3a,8,8a-hexahydro-1,3a,8-trimethylpyrrolo[2,3-b]indol- 5-ol[8-(cis2,-6-dimethyl-morpholin-4-yl)octyl]-carbamate L-bitartrate hydrate; Mediolanum Farmaceutici, Milan, Italy] is a pseudo-reversible carbamate-type cholinesterase inhibitor (ChEI) which interacts with the catalytic and regulatory anionic site of the enzyme. Its effects on extracellular levels of acetylcholine (ACh), norepinephrine (NE), dopamine (DA), and serotonin (5-HT, 5-hydroxytryptamine) were studied in rat cortex by using a microdialysis technique coupled with high-performance liquid chromatography-electrochemical detection (HPLC-ECD). Conscious, freely moving rats were systemically [per os (p.o.) and subcutaneously (s.c.)] administered MF-268 with no ChEI in the probe. Cholinesterase inhibition in brain was assayed in parallel experiments. Oral administration of MF-268 (0.5, 2.0, and 5.0 mg/kg) produced a significant increase of extracellular ACh in cortex; the maximal increase was 300% [not significant (n.s.)], 460% and 1,200%, respectively. Maximal cholinesterase (ChE) inhibition was 2.3% (n.s.) at 9 hr and 9.7% (P < .05) at 12 hr after the 2.0 and 5.0 mg/kg doses, respectively. Norepinephrine and DA levels were increased 180% and 100% after the 5.0 mg/kg dose, respectively; 100% and 60% after the 2.0 mg/kg dose, respectively; and 70% for both amines after the 0.5 mg/kg dose, respectively. The elevation lasted at least 5 hr with the 2.0 and 5.0 mg/kg doses. There were no major changes in 5-HT levels at these three doses. Subcutaneous administration (0.5 and 2.0 mg/kg) produced a maximal 360% (5.5 hr) and 2,500% (5 hr) increase in extracellular ACh, respectively. Maximal ChE inhibition was 13% (0.5 mg/kg) and 41% (2.0 mg/kg). Neither 0.5 nor 2.0 mg/kg produced a consistent modification of NE. Only a transient increase in DA was seen with the 0.5 mg/kg dose. There were no changes in 5-HT levels at these two doses. MF-268-treated animals showed slight cholinergic side effects (chewing, tremor) at both doses. MF-268 administered intracortically through the microdialysis probe at a concentration of 50 microM induced a 5,900% increase in ACh levels at 6 hr. This effect started 30 min after injection and continued throughout the period of administration. MF-268 produced a significant decrease in NE levels (-44%) starting at 30 min, and a slight but significant increase in DA levels of 45% at 2.5 hr. A significant increase of 5-HT (58%) was also observed starting at 4 hr. Slight symptoms of cholinergic toxicity were observed during intracortical administration.

6-Aminoquinolones: a new class of quinolone antibacterials?

A series of quinolone- and 1,8-naphthyridone-3-carboxylic acids, designed by previous QSAR studies and characterized by an amino group at the C-6 position instead of the usual fluorine atom, were synthesized for the first time and evaluated for in vitro antibacterial activity. All of the synthesized compounds maintain good activity against Gram-negative bacteria (Pseudomonas aeruginosa excluded), and those compounds having a thiomorpholine group as the C-7 substituent also have good activity against Gram-positive bacteria. Some aspects of structure-activity relationships associated with the C-1, C-5, C-7, and C-8 substituents are also discussed. Derivatives 18g and 38g displayed the best activity with geometric mean MICs of 0.45 and 0.66-0.76 micrograms/mL against Gram-negative and Gram-positive bacteria, respectively. This antimicrobial activity reflects their ability to inhibit bacterial DNA-gyrase. The results of this study show that, while the C-6 fluorine is still the preferred substituent, good activity can still be obtained by replacing it with an amino group.

7-(Disubstituted thiazolyl)-3,5-dihydroxy-6-heptenoic/heptanoic acid derivatives as HMG-CoA reductase inhibitors.

A series of disubstituted thiazoles, functionalized with the essential 3,5-dihydroxy-6-heptenoic or heptanoic chain, were prepared and evaluated for their ability to inhibit the enzyme HMG-CoA reductase in vitro. All the synthesized compounds 46-61 showed a moderate inhibitory potency.

Quinolinecarboxylic acids. 3. Synthesis and antibacterial evaluation of 2-substituted-7-oxo-2,3-dihydro-7H-pyrido[1,2,3-de] [1,4]benzothiazine-6-carboxylic acids related to rufloxacin.

A series of 2-substituted-7-oxo-2,3-dihydro-7H-pyrido[1,2,3-de][1,4]benzothiazine-6 -carboxylic acids has been prepared and evaluated for in vitro antibacterial activity. These derivatives were less active than corresponding desmethylated analogues. Among these derivatives, the most active compound 22a was selected for preliminary pharmacokinetics in rats. The pharmacokinetic data indicated that 22a was rapidly absorbed and induced lasting plasma and urinary levels. In comparison with rufloxacin, it was excreted in low quantity in urine; a significant amount of desmethylated piperazinyl urinary metabolite was observed.

Determination of heptylphysostigmine in plasma by high-performance liquid chromatography with electrochemical detection.

Heptylphysostigmine is a new and very promising cholinergic drug for the treatment of Alzheimer disease. A method has been developed for its determination in plasma with a detection limit of 50 pg/ml. The drug was extracted in n-hexane by a simple one-step procedure, after buffering with sodium bicarbonate. Samples were analysed on a 25 cm x 4.6 mm I.D. silica column (5 microns particle size) using a mixture of acetonitrile, methanol and ammonium nitrate as mobile phase. Since this molecule is quite unstable in plasma, pyridostigmine bromide was added to samples to limit the decomposition. Physostigmine was employed as internal standard. The molecule was electrochemically detected by oxidizing potential (+0.75 V). The method was applied to the analysis of blood samples taken from one healthy volunteer administered this drug. In the same subject the inhibition rate of acetylcholinesterase in plasma and red cells was also measured.

Synthesis of phenothiazine derivatives as potential inhibitors of phospholipase C.

In order to study the structure-activity relationships of phenothiazine derivatives inhibiting phosphatidylinositol-specific phospholipase C (PI-PLC), the synthesis of some phenothiazine amide, amine and ester derivatives was performed mainly by reacting 10H-phenothiazine-10-propanoyl chloride with some amines and alcohols; the resulting amides were reduced with borane to yield the corresponding amines. Starting from 2-chloro and 2-trifluoromethyl-10H-phenothiazine-10-propanoyl chloride two amides were synthesized. The inhibiting activity on PI-PLC from human platelets is reported.

Synthesis of some carbamates of myo-inositol.

The syntheses are described of the 1-O-carbamoyl (11), 1-O-carbamoyl-2-O-stearoyl (10), 1-O-(acetylcarbamoyl)-2-O-stearoyl (12), 1-O-(heptylcarbamoyl) (13), 2-O-(heptylcarbamoyl) (14) 1,2-di-O-(heptylcarbamoyl) (15), and 1-O-(octadecylcarbamoyl) (16) derivatives of myo-inositol. None of these compounds had significant activity against phospholipase C.

Reversal of cognitive impairment by heptyl physostigmine, a long-lasting cholinesterase inhibitor, in primates.

Cholinergic replacement therapy for Alzheimer's disease using existing cholinesterase inhibitors is compromised by short duration, meagre benefits restricted to subgroups of patients, and peripheral toxicity. Heptyl physostigmine is a lipophilic carbamate derivative of physostigmine. In rhesus monkeys, heptyl physostigmine (0.2-0.9 mg/kg i.m.) fully reversed a scopolamine-induced cognitive impairment. Following oral administration in squirrel monkeys, heptyl physostigmine (8 mg/kg) induced long-lasting hypothermia (greater than or equal to 4 h), a centrally-mediated cholinergic effect. Erythrocyte acetylcholinesterase activity was inhibited by 86% at the time of peak hypothermia (180 min). Clinical trials with heptyl physostigmine will enable a more rigorous evaluation of cholinomimetic therapy for dementia.

The behavioral effects of heptyl physostigmine, a new cholinesterase inhibitor, in tests of long-term and working memory in rodents.

We assessed the effects of heptyl physostigmine, a new cholinesterase inhibitor, in a mouse tail-flick (TF) test, a mouse and rat passive avoidance test, a rat conditioned suppression-of-drinking (CSD) test, a rat Random Interval (RI) response rate test and a rat delayed matching-to-position (DMTP) test. In the TF test, a dose of 8.0 mg/kg of heptyl induced a significant antinociceptive effect that was in excess of 75% of the maximum possible effect 300 minutes after administration. In the mouse passive avoidance test, a dose of 3.0 mg/kg of heptyl fully reversed, and a dose of 1.0 mg/kg partially reversed, a scopolamine-induced (0.2 mg/kg) deficit. In the rat passive avoidance test, a dose of 8.0 mg/kg fully reversed a scopolamine-induced (0.2 mg/kg) deficit, while a dose of 4.0 mg/kg of heptyl was without effect. In the same experiment, a dose of 0.6 mg/kg of physostigmine partially reversed the scopolamine-induced deficit. In the CSD test, a dose of 8.0 mg/kg of heptyl fully reversed, and doses of 1.0 and 4.0 mg/kg of heptyl partially reversed, the deficit induced by scopolamine (0.4 mg/kg). In the RI response rate test, doses of 8.0 mg/kg and 0.6 mg/kg of physostigmine fully suppressed lever pressing for food rewards. Doses of 4.0 mg/kg of heptyl and below had no effect on lever-pressing rates. In the working memory test (DMTP), 4.0 mg/kg heptyl partially reversed the scopolamine-induced deficit (0.2 mg/kg) in the number of correct choices made, but did not affect the scopolamine-induced deficit in the number of trials completed.(ABSTRACT TRUNCATED AT 250 WORDS)

New analogs of physostigmine: alternative drugs for Alzheimer's disease?

The synthesis of a series of physostigmine analogs, in which the methylcarbamyl group has been substituted with monoalkylcarbamyl, dimethyl- and diethylcarbamyl groups, is reported. These compounds were prepared with the aim of investigating their possible therapeutic effects in the treatment of Alzheimer's type dementia. The new analogs of physostigmine are inhibitors of acetylcholinesterase from Electroforus electricus, with a value of the reactivation constant, k3 smaller than the one of physostigmine. The percentage of anticholinesterase activity in vitro and in vivo, the acute toxicity and some behavioural effects were also evaluated for selected derivatives. The reactivation constant, in vitro, supports the view that the derivatives described would be more suitable for therapeutic use than physostigmine.

Quinolonecarboxylic acids. 2. Synthesis and antibacterial evaluation of 7-oxo-2,3-dihydro-7H-pyrido[1,2,3-de][1,4]benzothiazine-6-carboxylic acids.

A series of pyridobenzothiazine acid derivatives was synthesized and their in vitro antibacterial activity was evaluated. The 1,4-benzothiazine intermediates, which by Gould-Jacobs quinoline synthesis produced pyridobenzothiazine acids, were prepared by hydrolytic basic cleavage of substituted 2-aminobenzothiazoles and successive cyclocondensation with 1-bromo-2-chloroethane or alternatively with monochloroacetic acid, hence reduction by LiAlH4. The pyridobenzothiazine acids 10c, 30, and 31 show potent antibacterial activities against Gram-positive and Gram-negative pathogens. Structure-activity relationships are discussed. The compound 9-fluoro-10-(4-methyl-1-piperazinyl)-7-oxo-2,3-dihydro-7H-pyrido[1,2,3-d e] [1,4]benzothiazine-6-carboxylic acid (31) (MF-934) has been found to possess, together with the antibacterial activity, a weak acute toxicity and interesting pharmacokinetic characteristics in several animal species (rat, dog, monkey, man).

A long-lasting cholinesterase inhibitor affecting neural and behavioral processes.

A series of analogues of physostigmine were prepared with the aim of investigating their inhibitory effects on acetylcholinesterase in the treatment of Alzheimer's disease. One of the isomers prepared was evaluated for its anticholinesterase activity in vivo, acute toxicity, and some behavioral effects. This compound was a competitive inhibitor of the enzyme and was found to antagonize the stimulating effect produced by scopolamine on locomotor activity and to facilitate memory consolidation.

Synthesis and beta-adrenergic blocking activity of oxipropanolamines of 3,4-dihydro-3-oxo-2H(1,4)benzothiazine.

The synthesis of a series of oxypropanolamines of 3,4-dihydro-3-oxo-2H(1,4)benzothiazine is reported. Some of these compounds proved more potent than propranolol and carteolol as beta-adrenergic blocking agents in in vitro tests. The 8-(3-tert-butylamino-2-hydroxy)propoxy-3,4-dihydro-3-oxo-2H(1,4) benzothiazine fumarate (XVI a), which gave better results, confirmed its remarkable activity in in vivo tests.