The present study was carried out in an attempt to make a gelatin film strongly bioadhesive by introducing free dangling aldehyde groups. When gelatin films were treated with 0.5M of glutaraldehyde (GA) solution at 60 degrees C, free aldehyde groups (up to 150 micromol/g) were introduced in the film. The bonding strength of GA-crosslinked gelatin films (GA gelatin films) with biological tissue was assessed using porcine skins. It was found that bonding strength increased with increasing aldehyde content in the film. The GA gelatin films had bonding strength as high as 250 gf/cm2 whereas the native gelatin film (before GA treatment) showed bonding strength of 40 gf/cm2. When the aldehyde groups introduced in the gelatin films were quenched with glycine or reduced by NaBH4, the films no longer demonstrated such high bonding strength. These facts suggest that a Schiff base was formed between the free dangling aldehyde in the GA gelatin films and the amino groups of the natural tissue, which strongly contributed to a marked bioadhesion.

Although fibrin glue has been clinically used as a surgical adhesive, hemostatic agent, and sealant, it has the risk of virus infection because its components, fibrinogen and thrombin, are obtained from human blood. To circumvent this problem, we employed bioabsorbable gelatin and polysaccharides to prepare a safer hemostatic glue. Gelatin was modified with ethylenediamine using water-soluble carbodiimide to introduce additional amino groups into the original gelatin, while dextran and hydroxyethyl-starch were oxidized by sodium periodate to convert 1,2-hydroxyl groups into dialdehyde groups. Upon mixing of the two polymer components in aqueous solution, Schiff base was formed between the amino groups in the modified gelatin and the aldehyde groups in the modified polysaccharides, which thus resulted in intermolecular cross-linking and gel formation. The fastest gel formation took place within 2 s, and its bonding strength to porcine skin was about 225 gf cm(-2) when 20 wt% of an amino-gelatin (55% amino) and 10 wt% of aldehyde-HES (>84% dialdehyde) aqueous solutions were mixed. In contrast, the gelation time and bonding strength of fibrin glue was 5 s and 120 gf cm(-2), respectively.

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Susceptibility to HIV infection was examined in macrophages differentiated from human monocytes by macrophage colony-stimulating factor (M-CSF) or granulocyte/macrophage colony-stimulating factor (GM-CSF). The replication of macrophage-tropic human immunodeficiency virus type-1 (HIV-1), which was determined by reverse transcriptase (RT) activity, was significantly suppressed in macrophages induced by GM-CSF (GM-type macrophages) but not in those induced by M-CSF (M-type macrophages). Multinucleated giant cells were formed only in M-type macrophages after HIV infection. However, the expression of CD4 molecules on the surface of both types of macrophages was similar and the proviral DNA was detectable in cell lysates of both macrophages, although the amount of proviral DNA in M-type macrophages was higher than that in GM-type macrophages. Many steps have been defined in HIV infection and replication, such as adsorption of HIV to the cell surface, internalization of the viral core into the cytoplasm, uncoating of viral RNA, reverse transcription and integration of proviral DNA into cellular DNA, transcription and translation of proviral DNA, assembly of viral components, and budding of virus particles. Our findings suggested that the suppression of HIV-1 replication in macrophages induced by GM-CSF is mainly due to a disturbance at certain steps of replication after synthesis of the proviral DNA. Thus, the suppression of HIV replication in GM-type macrophages may provide a model of the latency of HIV infection in vivo.

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The in vitro maturation of peripheral blood monocytes to macrophages can be followed morphologically, and by measurement of cell surface antigens (CD4, HLA-DR, and FcR III) and lysozyme production. We used these markers to correlate monocyte maturation with susceptibility to human immunodeficiency virus (HIV) infection. Maturation of peripheral blood monocytes is associated with a decrease in membrane CD4, while HLA-DR and FcR III expression increase along with lysozyme secretion. Cells at all stages of maturation were susceptible to HIV infection, even mature macrophages without CD4 detectably by immunofluorescent staining. Maximal replication was observed in 7-day-old cells.

We have developed a new biodegradable scaffold that does not require any cell seeding to create an in-situ tissue-engineering vasculature (iTEV). Animal experiments were conducted to test its characteristics and long-term efficacy. An 8-mm tubular biodegradable scaffold, consisting of polyglycolide knitted fibers and an L-lactide and ε-caprolactone copolymer sponge with outer glycolide and ε-caprolactone copolymer monofilament reinforcement, was implanted into the inferior vena cava (IVC) of 13 canines. All the animals remained alive without any major complications until euthanasia. The utility of the iTEV was evaluated from 1 to 24 months postoperatively. The elastic modulus of the iTEV determined by an intravascular ultrasound imaging system was about 90% of the native IVC after 1 month. Angiography of the iTEV after 2 years showed a well-formed vasculature without marked stenosis or thrombosis with a mean pressure gradient of 0.51±0.19 mmHg. The length of the iTEV at 2 years had increased by 0.48±0.15 cm compared with the length of the original scaffold (2–3 cm). Histological examinations revealed a well-formed vessel-like vasculature without calcification. Biochemical analyses showed no significant differences in the hydroxyproline, elastin, and calcium contents compared with the native IVC. We concluded that the findings shown above provide direct evidence that the new scaffold can be useful for cell-free tissue-engineering of vasculature. The long-term results revealed that the iTEV was of good quality and had adapted its shape to the needs of the living body. Therefore, this scaffold would be applicable for pediatric cardiovascular surgery involving biocompatible materials.

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