Oxygen‐Loaded Nanodroplets Effectively Abrogate Hypoxia Dysregulating Effects on Secretion of MMP‐9 and TIMP‐1 by Human Monocytes

Abstract

Monocytes play a key role in the inflammatory stage of the healing process. To allow monocyte migration to injured tissues, the balances between secreted matrix metalloproteinases (MMPs) and their inhibitors (TIMPs) must be finely modulated. However, a reduction of blood supply and local oxygen tension can modify the phenotype of immune cells. Intriguingly, hypoxia might be targeted by new effective oxygenating devices such as 2H,3H-decafluoropentane- (DFP-) based oxygen-loaded nanodroplets (OLNs). Here, hypoxia effects on gelatinase/TIMP release from human peripheral monocytes were investigated, and the therapeutic potential of dextran-shelled OLNs was evaluated. Normoxic monocytes constitutively released ~500 ng/mL MMP-9, ~1.3 ng/mL TIMP-1, and ~0.6 ng/mL TIMP-2 proteins. MMP-2 was not detected. After 24 hours, hypoxia significantly altered MMP-9/TIMP-1 balance by reducing MMP-9 and increasing TIMP-1, without affecting TIMP-2 secretion. Interestingly OLNs, not displaying toxicity to human monocytes after cell internalization, effectively counteracted hypoxia, restoring a normoxia-like MMP-9/TIMP-1 ratio. The action of OLNs was specifically dependent on time-sustained oxygen diffusion up to 24 h from their DFP-based core. Therefore, OLNs appear as innovative, nonconventional, cost-effective, and nontoxic therapeutic tools, to be potentially employed to restore the physiological invasive phenotype of immune cells in hypoxia-associated inflammation.

Dextran-shelled oxygen-loaded nanodroplets reestablish a normoxia-like pro-angiogenic phenotype and behavior in hypoxic human dermal microvascular endothelium

“…This evidence complements previous data on the uptake of OLNs by other eukaryotic cells, 553 including human keratinocytes [Prato et al, 2015 and monocytes 554 [Gulino et al, 2015]. OLNs did not display cytoxic effects on HMEC-1.…”

“…These effects were specifically dependent on time-sustained oxygen release 558 from the inner core of OLNs, since they were not reproduced after treatment with OFNs or 559 OSS. These results are in full agreement with those obtained from parallel works with 560 dextran-shelled OLNs, able to restore normoxia-like MMP-9/TIMP-1 ratio in hypoxic human 561 monocytes [Gulino et al, 2015], and chitosan-shelled OLNs, effective in abrogating hypoxia-562 dependent dysregulation of balances between gelatinases and their inhibitors in human 563 keratinocytes . Therefore, the findings proposed here appear extremely 564 relevant to reach a global vision of the pro-angiogenic phenotype of the chronic wound, since 565 endothelial cells play relevant roles during healing processes in concert with both monocytes 566 and keratinocytes [Eming et al, 2014].…”

“…To complement previous 488 data on hypoxia-dependent dysregulation of MMP/TIMP balances in human monocytes 489 [Gulino et al, 2015] and keratinocytes , the present in vitro study 490 aimed at investigating the effects of hypoxia on the pro-angiogenic phenotype and the wound 491 healing abilities of human dermal microvascular endothelial cells. Furthermore, innovative 492 and nonconventional dextran-shelled/DFP-cored OLNs were challenged for their potential 493 abilities to counteract the effects of hypoxia.…”

“…119 Therefore, it is extremely important to assess carefully the effects of hypoxia on each single 120 cell population participating to the wound healing process, from monocytes and keratinocytes 121 to endothelial cells and fibroblasts. In a couple of recent works published by our group 122 hypoxia was shown to reduce MMP-9 and increase TIMP-1 without affecting TIMP-2 123 secretion by human monocytes [Gulino et al, 2015], whereas in human keratinocytes 124 hypoxia was shown to reduce MMP-2, MMP-9, and TIMP-2 secretion without changing 125 TIMP-1 levels and venous ulcers). Topical oxygen therapy, based on an O 2 gas emulsion applied to the 143 superficial wound tissue, represents another promising approach to enhance the oxygenation 144 of wounded tisues [Sen, 2009].…”

Matrix metalloproteinases in the wound microenvironment: therapeutic perspectives

“…Indeed, chitosan-and dextran-shelled OLNs prevented detrimental effects of hypoxia on MMPs and TIMPs secretion and restored their balance in human keratinocytes and monocytes. 99,100 Furthermore, OLNs promoted proangiogenic phenotype in hypoxic human dermal endothelium, which can also result from restoration of appropriate MMP/ TIMP balance.…”