Microspheres of biodegradable polymers as a drug-delivery system in the vitreous.

Microspheres of biodegradable polymers were evaluated as a potential controlled-release drug-delivery system in the vitreous. The microspheres were prepared with polymers of poly(lactic acid) or copolymers of glycolic acid and lactic acid. The release of 5-fluorouracil (5-FU) from the microspheres was studied in vitro. Poly(lactic acid) microspheres released 70-85% of total 5-FU over 7 days. Microspheres of polymers with a smaller molecular weight released the drug more rapidly. Copolymer microspheres released 98% of 5-FU over 2 days. The rate of drug release was controllable by changing the molecular weight of the polymers or using a matrix of copolymer. The intravitreal kinetics of the microspheres were studied in ten rabbits in vivo. A suspension of microspheres was injected into the vitreous cavity of five normal eyes and five vitrectomized eyes. By 48 +/- 5.2 days after injection, the microspheres disappeared from the vitreous cavity in the five normal eyes. Clearance from the vitreous cavity was accelerated in the five rabbits that underwent vitrectomy (14 +/- 2.4 days; P less than 0.001). No difference was found in the b waves of electroretinograms before and after injection of the microspheres. The histologic study showed no abnormal findings as a result of the injection. These results suggested that microspheres of biodegradable polymers may be a potential delivery system for the controlled release of drugs in the vitreous.

Treatment of malignant pleural effusions with doxorubicin hydrochloride-containing poly(L-lactic acid) microspheres.

For the treatment of malignant pleural effusions, we prepared doxorubicin hydrochloride (Adriamycin)-containing poly(L-lactic acid) microspheres (ADR-MS). In vitro, 50 percent and 100 percent release times of ADR from ADR-MS were 6.3 and 20 days, respectively. After intrapleural administration of ADR-MS for seven patients at an ADR dose of 40 mg, ADR was detected in the effusions for more than two weeks; however, ADR concentrations in serum were very small, consistent with minimal transpleural absorption of ADR. These results indicated the slow release of ADR into the pleural cavity. Furthermore, the amount of drained ADR was less than a few percent of the administered dose. In some cases, malignant cells in the effusion disappeared after the treatment. No complications related to the procedure occurred, and the patients developed no systemic symptoms. One patient died after four months, and the other six patients are alive after 21 to 31 months without reaccumulation of the effusion. The local administration of ADR-MS produces a localized high and systemic low concentration of ADR, which could potentially improve the patient's quality of life.

Controlled cisplatin delivery system using poly(D,L-lactic acid).

Cisplatin (CDDP)-containing poly(D,L-lactic acid) microspheres (CDDP-MS) and beads (CDDP-B) with an average molecular weight of the oligomer of 1.2 x 10(4) and 4% CDDP loading were prepared. In Tris buffer, 95% of CDDP disappeared from CDDP-MS within 3 d. In vitro and in vivo, CDDP-B released CDDP for 30-57 d, and for 21-42 d, respectively. The other CDDP-B with an average oligomer molecular weight of 9.6 x 10(3) with 5% lactic acid monomers, that contained 4% CDDP, showed a two-phase CDDP release pattern and CDDP disappeared within 41 d in vitro, and within 21 d in vivo. Histologically, tissue necrosis surrounding the CDDP-B was not severe.

Evaluation of a bilayer artificial skin capable of sustained release of an antibiotic.

A bilayer artificial skin, composed of an upper silicone sheet and a lower collagen sponge, has been developed by modifying a technique proposed by Yannas and Burke. We have applied it clinically with success, but infection sometimes occurred in the area where the artificial skin was placed. To use it safely in an infected wound, we developed a new type of artificial skin capable of sustained release of antibiotic. Microspheres of poly-L-lactic acid containing an antibiotic, were installed in the upper silicone sheet. The usefulness of the new type of artificial skin was suggested by in vitro studies.

Biodegradation and antitumour effect of adriamycin-containing poly(L-lactic acid) microspheres.

Adriamycin-containing poly (L-lactic acid) microspheres were prepared to develop a slow-releasing and long-acting adriamycin delivery system. An almost constant release of adriamycin from the adriamycin-containing poly(L-lactic acid) was achieved in Tris buffer and adriamycin disappeared within 20 d. Adriamycin was not detected in serum for up to 14 d, when the suspension of the adriamycin-containing poly(L-lactic acid) microspheres was injected into lung parenchyma, the femoral muscles of rabbits or the peritoneal cavity of mice. However, adriamycin remained in the rabbit muscles for up to 10 d under formation of scar tissue. When free adriamycin was added to P815 tumour cells in culture, the cell survival rate decreased with the exposure time. The treatment with the adriamycin-containing poly(L-lactic acid) microspheres showed a higher survival rate for mice bearing P815 tumour cells than with free adriamycin. In addition, the systemic side effects were insignificant when the adriamycin-containing poly(L-lactic acid) microspheres were given to mice instead of free adriamycin.

beta-Lactoglobulin A with N-ethylmaleimide-modified sulfhydryl residue, polymerized through intermolecular disulfide bridge on heating in the presence of dithiothreitol.

Roles of sulfhydryl groups on thermal aggregation of beta-lactoglobulin A (betaLG A) at pH 7.5 were investigated. It is known that betaLG A modified at Cys(121) with N-ethylmaleimide (NEM-betaLG A) does not form an aggregate by heating and that dithiothreitol (DTT) reduces cystine residues and induces the intermolecular sulfhydryl/disulfide interchange reaction and/or oxidation. NEM-betaLG A was heated in the presence of DTT. The molecules were linked together with an intermolecular disulfide bridge, and the polymer formed increased with increase in DTT concentration. The largest portion of polymer was formed when DTT was added at around the same molar concentration as that of NEM-betaLG A. Then, polymer formation decreased with further increase in DTT concentration. The results suggest that sulfhydryl/disulfide residues other than Cys(121), generated from cysteine residues, can induce intermolecular sulfhydryl/disulfide interchange reactions to polymer and that thiol compounds, for example, added DTT, are capable of starting such reactions.

A bioabsorbable delivery system for antibiotic treatment of osteomyelitis. The use of lactic acid oligomer as a carrier.

We prepared a composite of D,L-lactic acid oligomer and dideoxykanamycin B for use as a biodegradable antibiotic delivery system with sustained effect. The composite was implanted in the distal portion of the rabbit femur, and the effective concentration of the antibiotic was measured in the cortex, the cancellous bone, and the bone marrow. In all bone tissues around the implant, the concentration of antibiotic exceeded the minimum inhibitory concentration for the common causative organisms of osteomyelitis for six weeks. Most of the implant material had been absorbed and the bone marrow had been repaired to a nearly normal state within nine weeks of implantation. The implant caused no systemic side effects, and it is likely to prove clinically useful as a drug delivery system for treating chronic osteomyelitis.

In vitro detection of equine arteritis virus from seminal plasma for identification of carrier stallions.

Equine arteritis virus (EAV) was readily isolated in RK-13 cell monolayers by plaque assay from seminal plasma of experimental carrier stallions when they contained high titers of virus regardless of the presence of non-viral cytotoxicity in the seminal plasma. The cytotoxicity interfered with virus isolation from seminal plasma which contained virus at titers less than 10 PFU/ml. However, it was possible to detect the virus in seminal plasma pretreated with PEG (#6000). EAV was consistently identified by RT-PCR from crude seminal plasma which contained virus at titers of more than 10(2.7) PFU/ml. In vitro detection of EAV by virus isolation supplemented with RT-PCR using seminal plasma was proved to be an effective alternative to the standard test mating as a diagnostic method for carrier stallions.

Macroscopic anatomy of the auditory tube diverticulum (guttural pouch) in the thoroughbred equine--a silicon mold approach.

The characteristics of the equine auditory tube diverticulum (guttural pouches) were studied in detail by the naked eye and silicon mold method. First, we examined the anatomical relationship between the guttural pouches and their associated bones and muscles. Secondly, a silicon mold was fabricated to clarify the three-dimensional aspect of the guttural pouches, paying a special attention to the distribution of major arteries and nerves surrounding it. Thirdly, the volume of the silicon mold was measured by immersing it in water. The guttural pouches are a pair of pouches located dorso-posteriorly to the posterior pharynx. The pouches had a close contact rostrally with the Os sphenoidale, ventrally with the pharynx and the esophagus, and caudally with the Articulatio atlantoaxialis. The left and right guttural pouches had almost the same capacity in each horse. The A. carotis interna, cranial ganglia of cervical nerves and cervical Truncus sympathicus, as well as the N. vagus, N. glossopharyngeus, N. hypoglossus, and Radices spinales of the N. accessorius, were present in the mucosal crease extending from the roof of the guttural pouches towards the middle of the caudal wall. The N. facialis appeared from the Foramen stylomastoideum ran dorsocaudally along the lateral recess. The N. mandibularis appeared from the Foramen lacerum went over the muscular process of an ear bone, ran along the roof of the lateral recess and finally continued with the rostral side of the guttural pouches. Lymph nodes, scattered on the mucosal surface, appeared as tiny indentations with approximately millet seed size on the silicon surface. The capacity of guttural pouches in adult horses (472 +/- 12.4 cm3) was three-fold larger than that in foals (145 +/- 9.4 cm3).

[Intravitreal drug delivery by microspheres of biodegradable polymers].

We evaluated the efficacy of microspheres of biodegradable polymers as a slow releasing drug delivery system in the vitreous body. Microspheres containing 5-FU were prepared with polymers of poly-(lactic acid) or copolymers of glycolic acid and lactic acid. The release of the drug was studied in vitro. Poly-(lactic acid) microspheres released 5-FU for 7 days. The intravitreal kinetics of the microspheres was studied in rabbits in vivo. The microspheres disappeared from the vitreous cavity of normal eyes by 48 +/- 5 days after injection. Disappearance was accelerated from the vitreous cavity of vitrectomized rabbits (14 +/- 2 days, p less than 10(-6)). No abnormality was found on electroretinographic or histological examinations after microspheres injection. These results suggested that microspheres of biodegradable polymers could be useful as a potential drug delivery system for sustained drug release in the vitreous body.

In vitro evaluation of sustained drug release from biodegradable elastomer.

Poly(DL-lactic acid) (PLA), poly(epsilon-caprolactone) (PCL), and their copolymers (PLA-CL) with various monomer compositions were synthesized, and their properties as matrix for the sustained release of drugs were evaluated. The copolymerization technique produced very soft films which incorporated the drugs without deterioration of the elastic properties. Cisplatin and MD-805 were loaded in the films by casting the polymer solution containing the drugs. Fractions of the drugs released from the PLA-CL films were governed by the initial loading, the film thickness, and the polymer molecular weight. The drug release profiles obeyed the classical Fickian diffusion equation at least in the early stage, but significant hydrolytic degradation of the matrix polymers occurred in the later stage, influencing the kinetics of drug release. The monomer composition of copolymer affected the release profile more strongly than the initial molecular weight of the copolymer.

New biodegradable oligoesters for pharmaceutical application.

Tartaric acid, malic acid, and glyceric acid were copolycondensed with glycolic acid at various molar ratios in feed to quickly synthesize biodegradable oligoesters. They were likely to have a moderately cross-linked structure with relatively low molecular weights and hydrophilic groups on the chains. In addition to macroscopic gels which were insoluble in any solvents, we could obtain the oligoesters which were insoluble in water but soluble in N,N-dimethylformamide. The degradation rate of the oligoesters was higher than that of lactic acid (LA) oligomers having molecular weights of a few thousands. On the contrary, their glass transition and flow temperatures were much higher than those of LA oligomers, indicating that their handling during the preparation of drug delivery dosage forms was much improved. The formulation of microspheres containing drugs from the oligoesters revealed that they were useful as biodegradable matrices having high degradation rates.

Biodegradability of oxidized poly(vinyl alcohol).

Poly(vinyl alcohol) dehydrogenase (PVADH) purified from Pseudomonas sp. 113P3 catalyzed an oxidation of poly(vinyl alcohol) (PVA) in the presence of pyrroloquinoline quinone (PQQ) to give a beta-diketone structure on PVA. Although PVADH oxidized not only enzymatically oxidized PVA but also chemically oxidized PVA, PVA-degrading microorganisms, Pseudomonas sp. 113P3 and Arthrobacter tumescens sp. 52-1 grew on the enzymatically oxidized PVA, but not on the chemically oxidized PVA. This suggests that the growth of PVA-degrading microorganisms is affected by the structure of oxidized PVA.

Vero cell growth and differentiation on poly(L-lactic acid) membranes of different pore diameters.

In the last few years, the demand has increased for research on polymeric materials, which can be used as substitutes for injured tissues and organs or to improve their regeneration. In this work, we studied poly(L-lactic acid) (PLLA) membranes, a resorbable biomaterial, which were either dense or had different pore diameters (less than 45 microm, between 180 and 250 microm, and between 250 and 350 microm), in relation to stimulation of cell adhesion, growth, and differentiation in vitro. We used Vero cells, a fibroblastic cell line, as the biological model of investigation. We found that cells attached slowly to all PLLA membranes studied. On the other hand, once the adhesion occurs, the cells are able to grow and differentiate on the different polymers. The cells grew to form a confluent monolayer and were capable of producing collagen Type IV and fibronectin on different PLLA membranes. This behavior indicates that cells try to create a better environment to stimulate their growth. This also indicates that Vero cells alter their differentiation pattern once they are producing extracellular matrix molecules related to epithelial differentiation.

Determinação da maturidade esquelética e estimativa da idade através de radiografias carpais / Determination of skeletal maturity and estimante of the age through carpal radiographics

O autor, neste trabalho, tem como objetivo a determinação de um modelo para estimar a idade de indivíduos a partir da área de capeamento epifisário do rádio observada em radiografias de mão e punho esquerdo. As áreas do capeamento epifisário do rádio foram determinadas através da utilização de um aplicativo com base em imagens digitalizadas em computador. Foram utilizadas 55 radiografias de indivíduos do sexo masculino e 55 do sexo feminino, com idade variando entre sete e dezoito anos. Foram obtidos modelos para indivíduos do sexo feminino e sexo masculino. A análise dos dados mostram que o aumento da idade está associada à diminuição da área de capeamento epifisário do rádio. Os resultados deste estudo evidenciam a conveniência e o potencial quanto ao uso de modelos de regressão, na obtenção de estimativas da idade dos indivíduos em função da área de capeamento epifisário do rádio