Synthesis, characterization, biodegradation, and drug delivery application of biodegradable lactic/glycolic acid polymers: Part III. Drug delivery application

Lactic/glycolic acid polymers (PLGA) are widely used for drug delivery systems. The microsphere formulation is the most interesting dosage form of the PLGA-based controlled release devices. In this study, the previously reported PLGA were used to prepare drug-containing microspheres. Progesterone was used as a model drug. The progesterone microspheres were prepared from PLGA having varied compositions and varied molecular weight. The microscopic characterization shows that the microspheres are spherical, nonaggregated particles. The progesterone-containing PLGA microspheres possess a Gaussian size distribution, having average size from 70-134 microm. A solvent extraction method was employed to prepare the microspheres. The microencapsulation method used in this study has high drug encapsulation efficiency. The progesterone release from the PLGA microspheres and the factors affecting the drug release were studied. The release of progesterone from the PLGA microspheres is affected by the properties of the polymer used. The drug release is more rapid from the microspheres prepared using the PLGA having higher fraction of glycolic acid moiety. The drug release from the microspheres composed of higher molecular weight PLGA is faster. The drug content in microspheres also has an effect on the drug release. Higher progesterone content in microspheres yields a quicker initial burst release of the drug.     

Polymorphous low grade adenocarcinoma of minor salivary gland: a clinicopathologic and comparative immunohistochemical study

Retrospective review of seven cases of polymorphous low grade adenocarcinoma (PLGA) revealed that the original histologic diagnosis in five instances was mixed tumor (four benign and one malignant). Comparison of PLGA tumors with eleven representative cases of benign pleomorphic adenoma of minor salivary gland (BPA) showed marked similarity in cytologic features. PLGA and BPA also exhibited similar mucohyaline and/or fibromyxoid stromal components. In contrast, tumor margins of the two neoplasms were quite different; margins of PLGA were consistently infiltrative, whereas those of BPA were generally well circumscribed and encapsulated. Immunophenotypic comparison confirmed the glial fibrillary acidic protein (GFAP) positivity in BPA reported by others, whereas cases of PLGA were uniformly nonreactive with this antibody. Although previous reports have emphasized the histologic similarities of PLGA and adenoid cystic carcinoma, our findings indicate that differentiation of PLGA from BPA may represent a more common diagnostic problem. Awareness of comparative histologic features is important in the differential diagnosis of these minor salivary gland neoplasms. Also, monoclonal GFAP may be useful in distinguishing cases of PLGA from BPA where histologic features are otherwise similar.     

Curcumin-loaded into PLGA nanoparticles

The incorporation of the curcumin into poly(lactic-co-glycolic)acid (PLGA) nanospheres by the nanoprecipitation technique, the characterization of the nanoparticles and the schistosomicidal activity of the curcumin-loaded into PLGA nanospheres were reported. The incorporation process occurred with high efficiency and the images of field-emission scanning electron microscopy (FESEM) revealed the production of spherically shaped particles. According to the dynamic light scattering measurements, the particles are nanometric and monodisperse. The curcumin-loaded PLGA nanoparticles (50 and 100 μM) caused the death of all worms and a separation between 50% and 100% of Schistosoma mansoni couples at concentrations from 30 μM. Moreover, the curcumin-loaded PLGA nanoparticles also decreased the motor activity and caused partial alterations in the tegument of adult worms. This study marks the first time that schistosomicidal activity has been reported for curcumin-loaded PLGA nanoparticles.     

C-kit expression distinguishes salivary gland adenoid cystic carcinoma from polymorphous low-grade adenocarcinoma.

Adenoid cystic carcinoma (ACC) is characterized by persistent, relentless growth and a high rate of eventual metastasis. In contrast, polymorphous low-grade adenocarcinoma (PLGA) has a much lower risk of recurrence and rarely metastasizes. The histologic patterns of these two neoplasms can be similar. Expression of c-kit, a transmembrane receptor tyrosine kinase, has recently been reported to be expressed in ACC but not PLGA. Expression of galectin-3, a nonintegrin beta-galactosidase-binding lectin, has been reported to be significant in PLGA and decreased in ACC.Formalin-fixed paraffin-embedded tissue from 9 ACC and 14 PLGA were immunostained for c-kit and galectin-3. Cases were scored as 1+ (5-25% positive), 2+ (26-50% positive), or 3+ (>50% positive). C-kit was expressed by 100% of ACC (3+: 7 cases; 2+: 1 case; 1+: 1 case) and by 57% of PLGA (2+: 2 cases; 1+: 6 cases). In all but one ACC, c-kit expression was confined to the inner cell layer. C-kit expression was also noted in the intercalated duct epithelium of the salivary glands and the acinar cells of the lacrimal gland. Galectin-3 was expressed in 8 of 9 cases of ACC and 14 of 14 cases of PLGA.The results of this, the first study to compare c-kit and galectin-3 expression in ACC and PLGA, suggest that c-kit expression characterizes ACC, but not PLGA. Galectin-3 immunohistochemistry does not have a role in the differentiation of ACC and PLGA. C-kit immunostaining may be a valuable adjunctive tool for this differential diagnosis, particularly in the setting of a limited biopsy. Our finding of different patterns of c-kit expression in tubular and solid variants of ACC supports the concept of solid variant ACC as a high-grade tumor, with progression toward an entirely "inner cell" phenotype.     

Primary polymorphous low-grade adenocarcinoma of lung treated by sleeve bronchial resection: a case report

We report a surgical case of primary polymorphous low-grade adenocarcinoma (PLGA) of the minor salivary gland-type of the lung. A PLGA originating from the right upper lobar bronchial inlet was successfully treated by sleeve right upper lobectomy. PLGAs are thought to be indolent tumors that are preferentially localized to the palate, and they affect the minor salivary glands almost exclusively. Until now, two cases of distant metastases to the lung have been reported in the English literature. To the best of our knowledge, only one case of PLGA of minor salivary gland-type of the lung without evidence of a previous oropharyngeal primary tumor has been reported in the English literature. But the case was not a single lesion; it was bilateral tumors accompanied by tumors of the cervical lymph nodes. We report here the first case of a single primary PLGA of the minor salivary gland-type of the lung, which was successfully treated by sleeve bronchial resection of right upper lobe.     

Stabilizer-free poly(lactide-co-glycolide) nanoparticles for multimodal biomedical probes

Apart from the reported PLGA submicro- and microspheres with broad size distribution, we have successfully developed a methodology using nanoprecipitation to prepare different sizes of PLGA nanoparticles with narrow size distributions. The newly developed PLGA nanoparticles could be readily modified with hydrophilic biomaterials on their surface and entrap hydrophobic drugs into their interiors. The encapsulation of FITC inside PLGA nanoparticles displayed a controlled release of drug system. The surfaces of the FITC entrapped PLGA nanoparticles were conjugated with quantum dots to serve as bimodal imaging probes. For nuclear transport, combination of nuclear localization signal (NLS) and PLGA nanoparticles, PLGA nanoparticles could successfully enter into HeLa cells nuclei. From tissue uptake results, PLGA nanoparticles had more uptaken by brain and liver than other tissues. The iron oxide nanoparticles-conjugated PLGA nanoparticle showed high efficiency of relaxivities r2 and could be used as the powerful magnetic resonance imaging (MRI) agents.     

Cytogenetic similarities between two types of salivary gland carcinomas: adenoid cystic carcinoma and polymorphous low-grade adenocarcinoma.

Adenoid cystic carcinoma (ACC) and polymorphous low-grade adenocarcinoma (PLGA) are low-grade adenocarcinomas of salivary glands with a putative common histogenesis from the intercalated ducts but featuring distinct histological appearances. Hybrid tumors containing areas with histological patterns of both neoplasms have been reported but, to our knowledge, the question of their genotypic similarity has not yet been approached. As part of an ongoing study on cytogenetic characterization of salivary gland tumors, from a group of 24 malignant neoplasms, three out of five cases of ACC and three of four cases of PLGA were selected for their similar karyotypic changes. All of them displayed chromosome 12 abnormalities, affecting the 12q12-q13 region in four (all ACC cases and one PLGA case), 12q22 in one PLGA case, and 12p12.3 in the remaining. From this group of neoplasms, one PLGA and one ACC showed the same t(6;12)(p21;q13). Our findings favor the concept that tumors of salivary glands displaying epithelial and myoepithelial phenotypes share a common histogenesis.     

聚乳酸乙醇酸与猕猴骨髓基质细胞的生物相容性

背景：聚乳酸乙醇酸与不同细胞的生物相容性的报道较多,但探讨其与猕猴骨髓基质细胞生物相容性的研究极少。 目的：观察猕猴骨髓基质细胞的生物学特性以及其与支架材料聚乳酸乙醇酸在体外的生物相容性。 方法：将经流式细胞检测证实的第2代猕猴骨髓基质细胞与聚乳酸乙醇酸支架材料联合培养,以单独培养的骨髓基质细胞作为对照。 结果与结论：猕猴骨髓基质细胞与聚乳酸乙醇酸共培养4 d后,激光共聚焦显微镜下可见CD29阳性的骨髓基质细胞黏附包绕在聚乳酸乙醇酸纤维表面,形成长的细胞链。扫描电镜下可见骨髓基质细胞于聚乳酸乙醇酸上贴附良好,且有较多细长的突起从骨髓基质细胞伸出,沿材料伸展。骨髓基质细胞在聚乳酸乙醇酸浸出液中培养72 h,骨髓基质细胞神经营养因子表达量与对照组比较差异无显著性意义(P > 0.05);培养7 d,其形态、细胞活力、细胞增殖指数与对照组亦无明显差异。提示猕猴骨髓基质细胞与聚乳酸乙醇酸支架材料有良好的生物相容性。     

Delivering neuroactive molecules from biodegradable microspheres for application in central nervous system disorders

Nerve growth factor (NGF) may enhance axonal regeneration following injury to the central nervous system (CNS), such as after spinal cord injury. The release profile of NGF, co-encapsulated with ovalbumin, was tailored from biodegradable polymeric microspheres using both polymer degradation and protein loading. Biodegradable polymeric microspheres were prepared from PLGA 50/50, PLGA 85/15, PCL and a blend of PCL/PLGA 50/50 (1:1, w/w), where the latter was used to further tailor the degradation rate. The amount of protein loaded in the microspheres was varied, with PCL encapsulating the greatest amount of protein and PLGA 50/50 encapsulating the least. A two-phase release profile was observed for all polymers where the first phase resulted from release of surface proteins and the second phase resulted predominantly from polymer degradation. Polymer degradation influenced the release profile most notably from PLGA 50/50 and PLGA 85/15 microspheres. The amount and bioactivity of released NGF was followed over a 91 d period using a NGF-ELISA and PC12 cells, respectively. NGF was found to be bioactive for 91 d, which is longer than previously reported.     

Polymorphous low grade adenocarcinoma with distant metastases and deletions on chromosome 6q23-qter and 11q23-qter: a case report

Polymorphous low grade adenocarcinomas (PLGAs) are thought to be indolent tumours that are localised preferentially to the palate and affect the minor salivary glands almost exclusively. Metastases to locoregional lymph nodes occur in only 6-10% of cases. Recently, two cases of PLGA with microscopically confirmed distant metastases have been reported. This study reports a third case of PLGA with histologically and immunohistochemically confirmed distant metastases. It is the first case with multiple pleural, as well as pulmonary parenchymal, metastases and metastases in cervical and paraoesophageal lymph nodes. In most cases, PLGAs are salivary gland tumours with limited potential to metastasis and a good prognosis after local treatment. However, the recently reported cases reveal that the tumour can give rise to widely spread metastases. To obtain more information about the incidence of distant metastases, periodic chest x ray examination during follow up is desirable.     

Dual release of dexamethasone and TGF-β3 from polymeric microspheres for stem cell matrix accumulation in a rat disc degeneration model

Low back pain is frequently caused by nucleus pulposus (NP) degeneration. Tissue engineering is a powerful therapeutic strategy which could restore the normal biomechanical motion of the human spine. Previously we reported that a new nanostructured three-dimensional poly(lactide-co-glycolide) (PLGA) microsphere, which is loaded with dexamethasone and growth factor embedded heparin/poly(l-lysine) nanoparticles via a layer-by-layer system, was an effective cell carrier in vitro for NP tissue engineering. This study aimed to investigate whether the implantation of adipose-derived stem cell (ADSC)-seeded PLGA microspheres into the rat intervertebral disc could regenerate the degenerated disc. Changes in disc height by plain radiograph, T2-weighted signal intensity in magnetic resonance imaging (MRI), histology, immunohistochemistry and matrix-associated gene expression were evaluated in normal controls (NCs) (without operations), a degeneration control (DC) group (with needle puncture, injected only with Dulbecco’s modified Eagle’s medium), a PLGA microspheres (PMs) treatment group (with needle puncture, PLGA microspheres only injection), and PLGA microspheres loaded with ADSCs treatment (PMA) group (with needle puncture, PLGA microspheres loaded with ADSC injection) for a 24-week period. The results showed that at 24 weeks post-transplantation, the PM and PMA groups regained disc height values of ～63% and 76% and MRI signal intensities of ～47% and 76%, respectively, compared to the NC group. Biochemistry, immunohistochemistry and gene expression analysis also indicated the restoration of proteoglycan accumulation in the discs of the PM and PMA groups. However, there was almost no restoration of proteoglycan accumulation in the discs of the DC group compared with the PM and PMA groups. Taken together, these data suggest that ADSC-seeded PLGA microspheres could partly regenerate the degenerated disc in vivo after implantation into the rat degenerative intervertebral disc.     

A novel time-varying poly lactic-co glycolic acid external sheath for vein grafts designed under physiological loading

Changes in dimensional and mechanical properties of degradable sheaths in poly lactic-co glycolic acid (PLGA) have been researched extensively. Composite PLGA having variable resorption rates in multiple layers under physiological loading has not been reported. Our novel design of a PLGA sheath is composed of 3 layers with different degradation rates (i.e., the innermost layer degrades the fastest, followed by the middle, while the outer layer degrades the slowest). In the presence of physiological luminal pressure, diameter is greater, thickness is less, resorption rate is greater, pore size is greater, and incremental modulus is greater than in nonpressurized sheaths. Furthermore, the ratio of the pore size to the sheath radius affects the dimensional changes of the sheath in the radial direction. In addition to changing the pore size-to-sheath radius ratio, the dimensional changes can be manipulated by choosing different glycolic and lactic acid ratios for the different layers. The application of this novel PLGA design for gradual arterialization of vein grafts is contemplated.     

Reduction of inflammatory reaction of poly(d,l-lactic-co-glycolic Acid) using demineralized bone particles

Poly(lactide-co-glycolic acid) (PLGA) has been widely applied to tissue engineering as a good biocompatible material because of its biodegradability and nontoxic metabolites, but how the inflammatory reaction of PLGA on the surrounding tissue in vivo is reduced has not been discussed sufficiently. We hypothesized that the cells neighboring the PLGA implant might have an inflammatory response that could be reduced by impregnating demineralized bone particles (DBPs) into the PLGA. We manufactured five different ratios of DBP/PLGA hybrid materials, with each material containing 0, 10, 20, 40, and 80 wt% of DBPs of PLGA. For biocompatibility test, NIH/3T3 mouse fibroblasts were cultured on the DBP/PLGA scaffold for 3 days. The inflammatory potential of PLGA was evaluated using messenger ribonucleic acid expression of tumor necrosis factor alpha (TNF-alpha) and interleukin 1-beta (IL-1beta) on a human acute promyelocytic leukemic cell (HL-60). The in vivo response of DBP/PLGA film was compared with that of PLGA film implanted subcutaneously; the local inflammatory response was observed according to histology. The DBP/PLGA scaffold had no adverse effect on NIH/3T3 initial cell attachment and did not affect cell viability. DBP/PLGA films, especially PLGA films containing 80% DBP, elicited a significantly lower expression of IL-1beta and TNF-alpha from HL-60 cells than PLGA film alone. In vivo, DBP/PLGA film demonstrated a more favorable tissue response profile than PLGA film, with significantly less inflammation and fibrous capsule formation as below only 20% of DBP in PLGA film during implantation. This study shows that application of DBPs reduces the fibrous tissue encapsulation and foreign body giant cell response that commonly occurs at the interface of PLGA.     

Long-term variations in mechanical properties and in vivo degradability of CPC/PLGA composite

A new type of bone cement composite was successfully achieved by mixing degradable biosecure polylactic-co-glycolic acid (PLGA) fibers with high initial strength calcium phosphate cement (CPC). Its higher initial strength was mainly responsible for the in situ reinforcing effect of residual tetra-calcium phosphate monoxide (r-TTCP) particles reported in our previous work. So this bone cement composite containing fibers and the controlling group could be termed as CPC/PLGA composite and pure CPC or fiber-free group, respectively. In this study, we had investigated mechanical properties and microstructures of the CPC/PLGA composite immersed in 0.9% saline solution for different time and its in vivo degradation behaviors after implanting in rabbit muscle and femur bone, respectively. Results showed that the incorporation of the degradable fibers not only greatly increased the initial toughness and flexural strength of the CPC/PLGA composite but also significantly improved its later osteo-conduction as well as degradation rate. The rabbit muscle implant tests showed that the weight loss ratio of the CPC/PLGA composite increased by 41.03% as compared to the pure CPC. And the rabbit femur implant tests showed that the composite exhibits outstanding biocompatibility and bioactivity and more excellent osteoconduction and degradability than the pure CPC.     

Cell affinity for bFGF immobilized heparin-containing poly(lactide-co-glycolide) scaffolds

In order to effectively and uniformly immobilize basic fibroblast growth factor (bFGF) to thick PLGA scaffold, the heparin-conjugated PLGA (H-PLGA) was synthesized at the first by reaction between heparin and a low molecular weight PLGA. Then heparin-containing PLGA (H-PLGA/PLGA) scaffold was fabricated by blending the H-PLGA with a high molecular weight PLGA. Finally, bFGF was immobilized on the H-PLGA/PLGA scaffold mainly by static electricity action between them. The effect of H-PLGA content on bFGF binding efficiency of the H-PLGA/PLGA scaffolds was investigated. It was found that bFGF binding efficiency increased with increasing H-PLGA content. The bound bFGF can release in vitro slowly from the H-PLGA/PLGA scaffolds and last over two weeks. The released bFGF has still preserved its bioactivity. The attachment and growth of mouse 3T3 fibroblasts on the H-PLGA/PLGA scaffolds were better than that on the PLGA scaffold, however bFGF immobilized H-PLGA/PLGA scaffolds showed much better cell affinity. Therefore, the method to use the H-PLGA/PLGA scaffold for immobilizing bFGF is not only effective for slow delivering bFGF with bioactivity, but also can be used for fabricating thick scaffold where bFGF could be combined and uniformly distributed.     

Macroporous biodegradable natural/synthetic hybrid scaffolds as small intestine submucosa impregnated poly(D,L-lactide-co-glycolide) for tissue-engineered bone

—Poly( D , L-lactide-co-glycolide) (PLGA), a biodegradable synthetic polymer, is widely used in a variety of tissue-engineered applications, including drug-delivery systems. However, the PLGA scaffolds, macroporous and three-dimensional structure, are difficult to cell attachment and in-growth due to surface hydrophobicity. In order to introduce in new bioactive functionality from porcine small intestine submucosa (SIS) as natural source for PLGA, we fabricated SIS-powder-impregnated PLGA (SIS/PLGA) hybrid scaffolds. Fabrication parameters, including ratios of SIS, PLGA and salt, were optimized to produce the desired macroporous foam. The scaffolds had a relatively homogeneous pore structure, good interconnected pores from the surface to core region and showed an average pore size in the range 69.23–105.82 μm and over 90% porosity. The SIS/PLGA scaffolds degraded with a rate depending on the contents of the SIS. After the fabrication of the SIS/PLGA hybrid scaffolds the wettability of the scaffold was greatly enhanced, resulting in uniform cell seeding and distribution. So, it was observed that BMSC attachment to the SIS/PLGA scaffolds increased gradually with increasing SIS contents. Scaffolds of PLGA alone and SIS/PLGA were implanted subcutaneously under dorsal skin of athymic nude mouse to observe the osteoconductivity. It was found from the result that the effects of the SIS/PLGA scaffolds on bone formation are stronger than control PLGA scaffolds. In summary, the SIS/PLGA scaffolds have osteoconductive effects to allow remodeling and replacement by osseous tissue.     

Poly(lactic-co-glycolic acid) enhances maturation of human monocyte-derived dendritic cells

Immature dendritic cells (iDCs) were derived from human peripheral blood monocytes, and treated with 75:25 poly(lactic-co-glycolic acid) (PLGA) microparticles (MPs) or film to assess the resultant dendritic cell (DC) maturation as compared to positive control of lipopolysaccharide (LPS) treatment for DC maturation or negative control of untreated iDCs. The effect of PLGA contact on DC maturation was examined as one possible explanation for the PLGA adjuvant effect we have observed in the enhancement of an immune response to codelivered model antigen, as adjuvants act through the maturation of DCs. Culturing iDCs with PLGA MPs or PLGA film resulted in morphology similar to that of LPS-matured DCs and the association, or possible internalization, of PLGA MPs. Furthermore, biomaterial-treated iDCs demonstrated an increase in MHC class II and costimulatory molecule expression compared to iDCs but to a lower level than that of LPS-matured DCs. Direct iDC contact with PLGA MPs was necessary for maturation. Immature DCs exposed to PLGA MPs were stimulatory of allogeneic T-cell proliferation, whereas cells exposed to PLGA film were not. Further, PLGA MPs supported a moderate delayed type hypersensitivity reaction in mice indicative of in vivo DC maturation. Taken together, these results suggest that PLGA is a DC maturation stimulus and that the form of the biomaterial may influence the extent of DC maturation.     

Macroporous biodegradable natural/synthetic hybrid scaffolds as small intestine submucosa impregnated poly(D,L-lactide-co-glycolide) for tissue-engineered bone

Poly(D,L-lactide-co-glycolide) (PLGA), a biodegradable synthetic polymer, is widely used in a variety of tissue-engineered applications, including drug-delivery systems. However, the PLGA scaffolds, macroporous and three-dimensional structure, are difficult to cell attachment and in-growth due to surface hydrophobicity. In order to introduce in new bioactive functionality from porcine small intestine submucosa (SIS) as natural source for PLGA, we fabricated SIS-powder-impregnated PLGA (SIS/PLGA) hybrid scaffolds. Fabrication parameters, including ratios of SIS, PLGA and salt, were optimized to produce the desired macroporous foam. The scaffolds had a relatively homogeneous pore structure, good interconnected pores from the surface to core region and showed an average pore size in the range 69.23-105.82 microm and over 90% porosity. The SIS/PLGA scaffolds degraded with a rate depending on the contents of the SIS. After the fabrication of the SIS/PLGA hybrid scaffolds the wettability of the scaffold was greatly enhanced, resulting in uniform cell seeding and distribution. So, it was observed that BMSC attachment to the SIS/PLGA scaffolds increased gradually with increasing SIS contents. Scaffolds of PLGA alone and SIS/PLGA were implanted subcutaneously under dorsal skin of athymic nude mouse to observe the osteoconductivity. It was found from the result that the effects of the SIS/PLGA scaffolds on bone formation are stronger than control PLGA scaffolds. In summary, the SIS/PLGA scaffolds have osteoconductive effects to allow remodeling and replacement by osseous tissue.