This study investigates the fluorogenic characteristics of the chitosan-genipin reaction for applications in microencapsulation research. Results showed that the chitosan-genipin reaction generated a colored and fluorescent product, with optimal excitation and emission wavelengths at 369 and 470 nm, respectively. Furthermore, it was found that reaction conditions affected the fluorescence intensity of the product. Mixture at the ratio of 4:1 (chitosan: genipin by weight) fluoresced the most. It also fluoresced stronger if the reaction occurred at higher temperature, with the intensity of 10.4 x 10(5) CPS at 37 degrees C, 5.9 x 10(5) CPS at 20 degrees C, and 2.5 x 10(5) CPS at 4 degrees C. As well, the fluorescence of the mixture developed gradually over time, attaining the emission maxima of 2.9 x 10(5), 7.6 x 10(5), and 10.0 x 10(5) CPS in 1, 6, and 18 h, respectively. Chitosan-coated alginate microcapsules were prepared without prior labeling, to which subsequent genipin treatment was applied in order to examine the potential of using genipin in microcapsule characterization. Chitosan bound to the alginate beads interacted with genipin, from which the resultant fluorescent signals allowed for clear visualization of the chitosan coating under confocal laser scanning microscopy. The relative fluorescence intensity across the chitosan membrane was found to be considerably higher than the controls (175 vs. 50). The membrane thickness measured was 29.2 +/- 7.3 microm. These findings demonstrate a convenient and effective way of characterizing chitosan-based microcapsules using genipin as a fluorogenic marker, a technique that will be useful in microcapsule research and other biomedical applications.
The genipin cross-linked alginate-chitosan (GCAC) microcapsule, composed of an alginate core and a genipin cross-linked chitosan membrane, was recently proposed for live cell encapsulation and other delivery applications. This article for the first time describes the details of the microcapsule membrane characterization using a noninvasive and in situ method without any physical or chemical modifications on the samples. Results showed that the cross-linking reaction generated the fluorescent chitosan-genipin conjugates. The cross-linked chitosan membrane was clearly visualized by confocal laser scanning microscopy (CLSM). A straightforward assessment on the membrane thickness and relative intensity was successfully achieved. CLSM studies showed that the shell-like cross-linked chitosan membranes of approximately 37 microm in thickness were formed surrounding the microcapsule. The reaction variables, including cross-linking temperature and time significantly affected the fluorescence intensity of the membranes. Elevating the cross-linking temperature from 4 to 37 degrees C drastically intensified the membrane fluorescence, suggesting the attainment of a high degree of cross-linking on the chitosan membrane. Extended cross-linking time altered the cross-linked membranes in modulation. Although genipin concentration and cross-linking time had little effects on the membrane thickness, cross-linking at higher temperatures tended to form relatively thinner membranes.
Age-related macular degeneration (AMD) is a blinding disease caused by multiple factors and is the primary cause of vision loss in the elderly. The morbidity of AMD increases every year. Currently, there is no effective treatment option for AMD. Intravitreal injection of antivascular endothelial growth factor (anti-VEGF) is currently the most widely used therapy, but it only aims at neovascularization, which is an intermediate pathological phenomenon of wet AMD, not at the etiological treatment. Anti-VEGF therapy can only temporarily delay the degeneration process of wet AMD, and AMD is easy to relapse after drug withdrawal. Therefore, it is urgent to deepen our understanding of the pathophysiological processes underlying AMD and to identify integrated or new strategies for AMD prevention and treatment. Recent studies have found that autophagy dysfunction in retinal pigment epithelial (RPE) cells, cellular senescence, and abnormal immune-inflammatory responses play key roles in the pathogenesis of AMD. For many age-related diseases, the main focus is currently the clearing of senescent cells (SNCs) as an antiaging treatment, thereby delaying diseases. However, in AMD, there is no relevant antiaging application. This review will discuss the pathogenesis of AMD and how interactions among RPE autophagy dysfunction, cellular senescence, and abnormal immune-inflammatory responses are involved in AMD, and it will summarize the three antiaging strategies that have been developed, with the aim of providing important information for the integrated prevention and treatment of AMD and laying the ground work for the application of antiaging strategies in AMD treatment.
Cholesterol is known to be a major risk factor for coronary heart disease (CHD). Current treatments for elevated blood cholesterol include dietary management, regular exercise, and drug therapy with fibrates, bile acid sequestrants, and statins. Such therapies, however, are often suboptimal and carry a risk for serious side effects. This study shows that microencapsulated Lactobacillus plantarum 80 (pCBH1) cells can efficiently break down and remove bile acids, and establishes a basis for their use in lowering blood serum cholesterol. Results show that microencapsulated LP80 (pCBH1) is able to effectively break down the conjugated bile acids glycodeoxycholic acid (GDCA) and taurodeoxycholic acid (TDCA) with bile salt hydrolase (BSH) activities of 0.19 and 0.08 µmol DCA/mg CDW/h respectively. This article also summarizes the physiological interrelationship between bile acids and cholesterol and predicts the oral doses of microencapsulated Lactobacillus plantarum 80 (pCBH1) cells required for lowering cholesterol.
Microencapsulation is an emerging technology in the development of bioartificial organs for drug, protein, and delivery systems. One of the advancements in establishing an appropriate membrane material for live cell and tissue encapsulation is the incorporation of poly(ethylene glycol) (PEG) to the widely studied alginate microcapsules. The current study investigates the properties of integrating PEG to microcapsules coated with poly-L-lysine (PLL) and chitosan as well as a novel microcapsule membrane which combines both PLL and chitosan. Results show that microcapsules containing PEG can support cell viability and protein secretion. The addition of PEG to PLL and chitosan-coated microcapsules improves the stability of microcapsules when exposed to a hypotonic solution. We also compared the novel microcapsule with two other previously used microcapsules including alginate-chitosan-PEG and alginate-PLL-PEG-alginate. Results show that all three membranes are capable of providing immunoprotection to the cells and have the potential for long-term storage at -80 degrees C. The novel membrane containing PEG, chitosan, and PLL, however, revealed the highest cell viability and mechanical strength when exposed to external rotational force, but it was unable to sustain osmotic pressure. The study revealed the potential of using PEG-incorporated alginate, chitosan, and PLL microcapsules for encapsulating live cells producing proteins and hormones for therapy.
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