Safety of poly (ethylene glycol)-coated perfluorodecalin-filled poly (lactide-co-glycolide) microcapsules following intravenous administration of high amounts in rats

Abstract

The host response against foreign materials designates the biocompatibility of intravenously administered microcapsules and thus, widely affects their potential for subsequent clinical use as artificial oxygen/drug carriers. Therefore, body distribution and systemic parameters, as well as markers of inflammation and indicators of organ damage were carefully evaluated after administration of short-chained poly (vinyl alcohol, (PVA)) solution or poly (ethylene glycol (PEG))-shielded perfluorodecalin-filled poly (d,l-lactide-co-glycolide, PFD-filled PLGA) microcapsules into Wistar rats. Whereas PVA infusion was well tolerated, all animals survived the selected dose of 1247 mg microcapsules/kg body weight but showed marked toxicity (increased enzyme activities, rising pro-inflammatory cytokines and complement factors) and developed a mild metabolic acidosis. The observed hypotension emerging immediately after start of capsule infusion was transient and mean arterial blood pressure restored to baseline within 70 min. Microcapsules accumulated in spleen and liver (but not in other organs) and partly occluded hepatic microcirculation reducing sinusoidal perfusion rate by about 20%. Intravenous infusion of high amounts of PFD-filled PLGA microcapsules was tolerated temporarily but associated with severe side effects such as hypotension and organ damage. Short-chained PVA displays excellent biocompatibility and thus, can be utilized as emulsifier for the preparation of drug carriers designed for intravenous use.

Keywords: ALAT, alanine aminotransferase; ANOVA, one-way analysis of variance; ASAT, aspartate aminotransferase; Artificial oxygen carriers; BE, base excess; Biocompatibility; Biodegradable microcapsules; C3, complement factor 3; C4a, complement factor 4a; CARPA, complement activation-related pseudoallergy; CK, creatine kinase; DAPI, 4',6-diamidin-2-phenylindol; FITC-dextran, fluorescein isothiocyanate-dextran 150,000; IFN-?, interferon-gamma; IL, interleukin; IVM, intravital microscopy; LDH, lactate dehydrogenase; MAP, mean arterial blood pressure; PEG, poly (ethylene glycol); PFD, perfluorodecalin; PLA); PLGA, poly (d,l-lactide-co-glycolide); PVA, poly (vinyl alcohol); Perfluorocarbon; Poly (lactic/glycolic) acid (PLGA; Poly (vinyl alcohol); TNF-a, tumor necrosis factor alpha; pO2, pCO2, oxygen and carbon dioxide partial pressures.

Graphical abstract

Fig. 1

Effect of capsule infusion on mean arterial blood pressure. PFD-filled PLGA microcapsules (1.5 µm), 0.25% PVA or 0.9% NaCl were infused for 30 min (light gray, 20 ml/kg body weight × h). The values plotted are mean ± SEM of 14-6 (capsules), 9-3 (PVA and NaCl) individual experiments, *p < 0.05 compared to NaCl group for time-matched data (0–90 min).

Fig. 2

Organ distribution of PFD-filled PLGA microcapsules and effect of capsule infusion on tissue damage. PFD-filled PLGA microcapsules (1.5 µm) were stained with Nile red and were allowed to circulate for about 40 min within the blood circulation, before organs were cryopreserved and frozen. Cryosections of spleen (A), liver (B) and lung (C) were prepared and nuclei were stained with DAPI (blue) for fluorescence microscopic analysis (×200 magnification). For assessment of the effect of capsule infusion on tissue damage, unstained PFD-filled PLGA microcapsules (1.5 µm) were infused for 30 min (20 ml/kg body weight × h). Microscopic assessment (×100 magnification) of spleen (D), liver (E) and lung tissue (F).

Fig. 3

Effect of capsule infusion on organ/tissue damage: ALAT (A), ASAT (B), LDH (C) and CK (D). PFD-filled PLGA microcapsules (1.5 µm), 0.25% PVA or 0.9% NaCl were infused for 30 min (light gray, 20 ml/kg body weight × h). The values plotted are mean ± SEM of 14-6 (capsules), 9-3 (PVA and NaCl) individual experiments, *p < 0.05 compared to NaCl group.

Fig. 4

Effect of capsule infusion on release of cytokines and complement factors. PFD-filled PLGA microcapsules (1.5 µm), 0.25% PVA or 0.9% NaCl were infused for 30 min (20 ml/kg body weight × h). The values plotted are mean ± SEM of 6-4 (capsules) or 3 (PVA and NaCl) individual experiments, *p < 0.05 compared to NaCl group. The detection limit of the used ELISA was 0.08 ng/ml. Note: logarithmic scale was used for Fig. 4E, F and H. C4a level in NaCl and PVA groups at all time-points measured were below detection limit.

Fig. 5

Effect of capsule infusion on hepatic microcirculation. IVM was performed to determine the number of perfused vessels of the hepatic microvascular system after intravenous administration (30 min, light gray, 20 ml/kg body weight × h) of either 0.9% NaCl (A), PFD-filled PLGA microcapsules (1.5 µm, B) or PLGA microspheres (1.5 µm, C). The values plotted are mean ± SEM of 6 individual experiments.