Lihui Weng

In vitro and in vivo evaluation of biodegradable embolic microspheres with tunable anticancer drug release

Natural polymer-derived materials have attracted increasing interest in the biomedical field. Polysaccharides have obvious advantages over other polymers employed for biomedical applications due to their exceptional biocompatibility and biodegradability. None of the spherical embolic agents used clinically is biodegradable. In the current study, microspheres prepared from chitosan and carboxymethyl cellulose (CMC) were investigated as a biodegradable embolic agent for arterial embolization applications. Aside from the enzymatic degradability of chitosan units, the cross-linking bonds in the matrix, Schiff bases, are susceptible to hydrolytic cleavage in aqueous conditions, which would overcome the possible shortage of enzymes inside the arteries. The size distribution, morphology, water retention capacity and degradability of the microspheres were found to be affected by the modification degree of CMC. An anticancer drug, doxorubicin, was successfully incorporated into these microspheres for local release and thus for killing cancerous cells. These microspheres demonstrated controllable degradation time, variable swelling and tunable drug release profiles. Co-culture with human umbilical vein endothelial cells revealed non-cytotoxic nature of these microspheres compared to monolayer control (P>0.95). In addition, a preliminary study on the in vivo degradation of the microspheres (100-300μm) was performed in a rabbit renal embolization model, which demonstrated that the microspheres were compatible with microcatheters for delivery, capable of occluding the arteries, and biodegradable inside arteries. These microspheres with biodegradability would be promising for embolization therapies.

Doxorubicin loading and eluting characteristics of bioresorbable hydrogel microspheres: In vitro study

Non-bioresorbable drug eluting microspheres are being increasingly used for the treatment of unresectable liver tumors, whereas bioresorbable microspheres have not received much attention. In this study, bioresorbable microspheres prepared from chitosan and carboxymethyl cellulose were loaded with doxorubicin (Doxo) via ion-exchange interactions with carboxylic groups in the microspheres. With a 25-40% decrease in the microsphere size depending on their size ranges, the microspheres could load a maximum of 0.3-0.7 mg Doxo/mg dry spheres. As confirmed by confocal microscopy, Doxo was mainly concentrated in the outer 20±5 μm surface layer of the microspheres. The loaded microspheres were stable in aqueous dispersions without aggregation for a prolonged period of time but degradable in a lysozyme solution. Furthermore, the loaded microspheres exhibited a noticeable pH-sensitive behavior with accelerated release of Doxo in acidic environment due to the protonation of carboxylic groups in the microspheres. Compared to commercial non-resorbable drug eluting beads, the loaded bioresorbable microspheres showed a sustained release manner in phosphate buffered saline (PBS). The release data were fitted to an empirical relationship, which reveals a non-Fickian transport mechanism (n=0.55-0.59). These results demonstrate that the bioresorbable microspheres are promising as attractive carriers for Doxo.

An in situ forming biodegradable hydrogel-based embolic agent for interventional therapies

We present here the characteristics of an in situ forming hydrogel prepared from carboxymethyl chitosan and oxidized carboxymethyl cellulose for interventional therapies. Gelation, owing to the formation of Schiff bases, occurred both with and without the presence of a radiographic contrast agent. The hydrogel exhibited a highly porous internal structure (pore diameter 17±4 μm), no cytotoxicity to human umbilical vein endothelial cells, hemocompatibility with human blood, and degradability in lysozyme solutions. Drug release from hydrogels loaded with a sclerosant, tetracycline, was measured at pH 7.4, 6 and 2 at 37°C. The results showed that tetracycline was more stable under acidic conditions, with a lower release rate observed at pH 6. An anticancer drug, doxorubicin, was loaded into the hydrogel and a cumulative release of 30% was observed over 78 h in phosphate-buffered saline at 37°C. Injection of the hydrogel precursor through a 5-F catheter into a fusiform aneurysm model was feasible, leading to complete filling of the aneurysmal sac, which was visualized by fluoroscopy. The levels of occlusion by hydrogel precursors (1.8% and 2.1%) and calibrated microspheres (100-300 μm) in a rabbit renal model were compared. Embolization with hydrogel precursors was performed without clogging and the hydrogel achieved effective occlusion in more distal arteries than calibrated microspheres. In conclusion, this hydrogel possesses promising characteristics potentially beneficial for a wide range of vascular intervention procedures that involve embolization and drug delivery.

Bioresorbable Hydrogel Microspheres for Transcatheter Embolization: Preparation and in Vitro Evaluation

PURPOSE To develop and evaluate a bioresorbable spherical material for embolization. MATERIALS AND METHODS New bioresorbable hydrogel microspheres were prepared from carboxymethyl cellulose and chitosan by using an inverse emulsion method. Size distribution of the microspheres was determined with a microscope, and the colorability was tested with Evans blue dye. The compressibility was examined with a texture analyzer. After sieving, the suspendability of the microspheres was tested in saline solution/contrast agent mixture in different ratios, and then the injectability was tested with microcatheters (lumen sizes of 0.0165, 0.019, and 0.027 inches). The in vitro degradability tests were performed in a lysozyme solution. Cell culture study of the microspheres was performed with human fibroblasts. RESULTS The microspheres exhibit diameters of 100-1,550 μm with a transparent appearance. Their fracture force can reach 0.58-0.88 N, and fracture deformation varies from approximately 70% to 95% of their original size. These microspheres can be colored by Evans blue dye, and uniform subgroups of microspheres can be readily obtained by sieving. Within 3 minutes, the microspheres form a stable suspension in a 4:6 contrast agent/saline solution mixture, which can be easily injected through microcatheters without aggregating or clogging. These microspheres are biodegradable, with degradation times varying from 2 weeks to 1 month, depending on their composition. Cell culture studies reveal no adverse effect on the growth of human fibroblasts in the presence of the microspheres. CONCLUSIONS A biodegradable and noncytotoxic microsphere was successfully prepared. It can be well suspended in the contrast solution and easily injected through a microcatheter.

Intra-articular treatment of knee osteoarthritis: from anti-inflammatories to products of regenerative medicine

ABSTRACT Objectives: Knee osteoarthritis (OA) is a debilitating condition that may ultimately require total knee arthroplasty (TKA). Non-operative treatments are bracing, oral analgesics, physical therapy, and intra-articular knee injection (IAKI). The objective of this paper is to provide a systematic literature review regarding intra-articular treatment of knee OA and insight into promising new products of regenerative medicine that may eventually have a substantial effect on treatment. Methods: A literature search was executed using Medline, Cochrane, and Embase with keywords “knee osteoarthritis” and “injection.” Specifically, 45 articles that discussed intra-articular knee injection using corticosteroids, hyaluronic acid, analgesics, local anesthetics, and newer products of regenerative medicine, such as platelet-rich plasma (PRP) and mesenchymal stem cells (MSC), were analyzed. Of these, eleven were level 1, three were level 2, twelve were level 3, two were level 4, and seventeen were level 5 evidence. Papers included animal models. Results: Local anesthetics have potential side effects and may only be effective for a few hours. Morphine and ketorolac may provide significant pain relief for 24 hours. Corticosteroids may give patients weeks to months of effective analgesia, but complications may occur, such as systemic hyperglycemia, septic arthritis, and joint degradation . Hyaluronic acid is a natural component of synovial fluid, but efficacy with respect to analgesia is controversial. Platelet-rich plasma formulations, autologous conditioned serum, autologous protein solution, and mesenchymal stem cell injections contain anti-inflammatory molecules and have been proposed to attenuate joint destruction or potentially remodel the joint. Conclusions: Currently, knee OA treatment does not address the progressively inflammatory environment of the joint. More investigation is needed regarding products of regenerative medicine, but they may ultimately have profound implications in the way knee OA is managed.

Calibrated Bioresorbable Microspheres as an Embolic Agent: An Experimental Study in a Rabbit Renal Model

PURPOSE To evaluate the time frame of resorption and tissue response of newly developed bioresorbable microspheres (BRMS) and vessel recanalization after renal embolization. MATERIALS AND METHODS Embolization of lower poles of kidneys of 20 adult rabbits was performed with BRMS (300-500 µm). Two rabbits were sacrificed immediately after embolization (day 0). Three rabbits were sacrificed after follow-up angiography at 3, 7, 10, 14, 21, and 30 days. The pathologic changes in the renal parenchyma, BRMS degradation, and vessel recanalization were evaluated histologically and angiographically. RESULTS Embolization procedures were successfully performed, and all animals survived without complication. Infarcts were observed in all kidneys that received embolization harvested after day 0. Moderate degradation of BRMS (score = 1.07 ± 0.06) was observed by day 3. Of BRMS, 95% were resorbed before day 10 with scant BRMS materials remaining in the arteries at later time points. Partial vessel recanalization was observed by angiography starting on day 3, whereas new capillary formation was first identified histologically on day 7. Vascular inflammation associated with BRMS consisted of acute, heterophilic infiltrate at earlier time points (day 3 to day 10); this was resolved with the resorption of BRMS. Inflammation and fibrosis within infarcted regions were consistent with progression of infarction. CONCLUSIONS BRMS were bioresorbable in vivo, and most BRMS were resorbed before day 10 with a mild tissue reaction. Vessel recanalization occurred secondary to the resorption of BRMS.

Calibrated bioresorbable microspheres: a preliminary study on the level of occlusion and arterial distribution in a rabbit kidney model.

PURPOSE To assess the level of occlusion and arterial distribution of calibrated bioresorbable microspheres (BRMS-I and BRMS-II) compared with tris-acryl gelatin microspheres (TGMS) after renal embolization. MATERIALS AND METHODS Six rabbits underwent renal embolization with 100-300 µm BRMS-I and TGMS; three rabbits received partial occlusion (group 1, n = 3), and three rabbits received total occlusion (group 2, n = 3). Four other rabbits received 100-300 µm BRMS-II (with higher cross-linking density than BRMS-I) in the left kidneys reaching total occlusion (group 3, n = 4). Coronal sections of the kidneys were histologically analyzed. Ease of injection, microsphere deformation, vessel sizes, and arterial distribution were assessed. RESULTS The injection of BRMS-I, BRMS-II, and TGMS through microcatheters went smoothly without any clogging. In group 1, BRMS identification was easier than TGMS. In group 2, both BRMS-I and TGMS were observed in all three arterial levels (interlobar, arcuate, and interlobular arteries) without a significant difference (P = .84). BRMS-I were not significantly different from TGMS in the mean diameter of vessels occluded (197 µm ± 23 vs 158 µm ± 21, P = .25) or the microsphere deformation (8.85% ± 0.53% vs 11.80% ± 0.64%, P = .071). In group 3, the arterial distribution of BRMS-II was significantly different from BRMS-I and TGMS (P < .0001). CONCLUSIONS In occluding arteries, 100-300 µm BRMS-I were not significantly different from 100-300 µm TGMS. Arterial distribution of BRMS can be influenced by their cross-linking density.

In vitro comparative study of drug loading and delivery properties of bioresorbable microspheres and LC bead

Drug loadable bioresorbable microspheres (BRMS) are specially designed for the treatment of hypervascular tumors through arterial embolization. These microspheres consist of carboxymethyl chitosan crosslinked with carboxymethyl cellulose, and are available at different size ranges varying from 50 to 900 µm in diameter. Similar to commercially available non-resorbable drug eluting microspheres, LC Bead® microspheres (LCB), BRMS were capable of loading more than 99 % of doxorubicin, an anticancer drug, from the solution within 2 h with highly similar kinetics (difference factor f1 = 0.36; similarity factor f2 = 97.99). Doxorubicin loaded BRMS exhibited the highest elution rate in the 30 % ethanol aqueous solution saturated with potassium chloride, and the elution time depended on the ratio between the amount of loaded BRMS and the volume of elution media. After injection through microcatheters, BRMS have a higher recovery rate of the microsphere weight than LCB (90.96 vs. 79.63 %, P = 0.026). Although loaded BRMS eluted more drug into the injection medium than loaded LCB (8.63 vs. 3.80 %, P = 0.0015), there was no significant difference in the drug delivery rate between BRMS and LCB (83.88 vs. 86.65 %, P = 0.504). This study compares the loading capability as well as the drug delivery rate of BRMS and a commercial product under a condition simulating a transcatheter arterial chemoembolization procedure and demonstrates the potential of drug loaded BRMS for the treatment of hypervascular tumors such as hepatocellular carcinoma.

Particle embolization: Factors affecting arterial distribution

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In vitro evaluation of sunitinib loaded bioresorbable microspheres for potential application in arterial chemoembolization

Drug-loadable bioresorbable microspheres (BRMS) are designed for treating hypervascular tumors through chemoembolization, thereby reducing systemic side effects via controllable local delivery. The present study investigated the degradation and loading capability of bioresorbable microspheres with an anti-angiogenic agent, sunitinib, and then evaluated the release profiles in different media (PBS, 10μg/mL and 4mg/mL lysozyme solutions), and tested catheter deliverability as well as potential antiangiogenic effects of the loaded microspheres. The dry weight of the BRMS showed a consistent decrease over the period of incubation in a 10μg/mL lysozyme solution with 61.3% mass remaining on day 21. Sunitinib was loaded efficiently onto the microspheres, with smaller sizes exhibiting a slightly faster loading and release rate. At 2h, the loading percentages were 99.28%, 97.95%, and 94.39% for 100-300, 300-500, and 500-700μm microspheres, respectively. At 8h, the percentage of drug released were 78.4±5.8%, 71.7±0.3%, and 67.0±2.9% for 100-300, 300-500, and 500-700μm microspheres under static medium conditions, respectively. Under replacing-medium conditions, the presence of 10μg/mL lysozyme slightly delayed the drug release while 4mg/mL lysozyme significantly facilitated the drug release from the microspheres as compared with PBS solution. Confocal imaging revealed an even distribution of sunitinib throughout the microspheres. Drug loaded microspheres were delivered through microcatheters smoothly without any clogging. Sunitinib retained its efficacy at reducing the viability of human endothelial cells after elution from the microspheres. Thus, these bioresorbable microspheres are promising for arterial chemoembolization.