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Elijah Evans
Elijah Evans

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Developed by Angela He, it's completely free and surprisingly well done. The theme is quite a mature one too, with it touching on suicide and self-harm. It also touches on romance, there's a few memes (who doesn't love a good meme) and so on. What's striking initially is the artwork, it's seriously good. Great chilled-out soundtrack to go along with it too, the quality here really is impressive.

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You can find it free on and Steam and donate if you enjoyed the experience. You can also see more games made by He on their itch page, I'm certainly interested in seeing something longer from them given how impressed I was with it.

Two years ago today we announced, a secure, fast, privacy-first DNS resolver free for anyone to use. In those two years, has grown beyond our wildest imagination. Today, we process more than 200 billion DNS requests per day making us the second largest public DNS resolver in the world behind only Google.

CORE Turkey, Turkey Meal & Duck recipe provides protein-rich, natural, grain-free nutrition. This nutrient-dense recipe has Turkey & Duck for cats who prefer a fish-free, chicken-free diet. To help support optimal health and digestion, cranberries and probiotics are added.

While considerable progress has been made towards understanding the complex processes and pathways that regulate human wound healing, regenerative medicine has been unable to develop therapies that coax the natural wound environment to heal scar-free. The inability to induce perfect skin regeneration stems partly from our limited understanding of how scar-free healing occurs in a natural setting. Here we have investigated the wound repair process in adult axolotls and demonstrate that they are capable of perfectly repairing full thickness excisional wounds made on the flank. In the context of mammalian wound repair, our findings reveal a substantial reduction in hemostasis, reduced neutrophil infiltration and a relatively long delay in production of new extracellular matrix (ECM) during scar-free healing. Additionally, we test the hypothesis that metamorphosis leads to scarring and instead show that terrestrial axolotls also heal scar-free, albeit at a slower rate. Analysis of newly forming dermal ECM suggests that low levels of fibronectin and high levels of tenascin-C promote regeneration in lieu of scarring. Lastly, a genetic analysis during wound healing comparing epidermis between aquatic and terrestrial axolotls suggests that matrix metalloproteinases may regulate the fibrotic response. Our findings outline a blueprint to understand the cellular and molecular mechanisms coordinating scar-free healing that will be useful towards elucidating new regenerative therapies targeting fibrosis and wound repair.

Our knowledge of the molecular and cellular events during mammalian tissue repair is extensive (see refs [1], [6], [7], [8]) and yet, even with such broad understanding of the wound repair process, regenerative medicine has failed to develop therapies that can perfectly regenerate skin. This stems partly from the dynamic reciprocity of cellular interactions and signaling pathways and partly from a lack of appropriate models to observe these interactions in a regenerative environment [9]. While wound repair in fetal mammals [10], [11], [12], [13], [14] and marsupials [15] has provided insight into the cellular and molecular regulation of scar-free healing, comparisons of wound repair between fetal mammals and adults has limitations, both biological and practical [16]. The developing fetus, at the time when it heals scar-free, has an immature endocrine system, is immuno-incompetent, is contained in a moist sterile environment, and its cells are in a state of chronic hypoxia [16]. Adult skin is more completely differentiated and adult wounds are open to desiccation and infection, two factors that seriously complicate wound repair. Other promising models of scar-free healing, such as the MRL mouse, which share the ability to regenerate ear punches with rabbits, hares, pikas, cows, pigs and cats [17], [18], [19] has proven less than perfect when challenged to heal excisional skin wounds [20], [21] casting doubt on the special regenerative powers of this inbred mouse model.

Given their seemingly absolute powers of regeneration, a recurring question has been whether wounds made outside of regenerating structures (e.g. limbs and tails) in adult urodeles are capable of scar-free healing or, like adult anurans, heal with a scar [29]. In this study we examined full thickness excisional (FTE) wound healing of dorsal back skin in adult axolotls. Using an established mammalian excisional wound model to directly characterize cutaneous wound healing in adult axolotls, we examined hemostasis, inflammation, new tissue formation and remodeling processes. Additionally, we induced metamorphosis in adult axolotls to test the hypothesis that loss of larval skin characters and transition to a terrestrial form results in fibrotic scarring following FTE flank wounds. Here we demonstrate that both aquatic and terrestrial axolotls are capable of perfect, scar-free skin regeneration. We discuss these findings in the context of mammalian wound repair and present a blueprint for investigating the cellular and molecular mechanisms that regulate scar-free skin healing in adult vertebrates.

Previous work in regenerating newt limbs suggested that reformation of the basement membrane (BM) facilitates dermal regeneration and its delayed formation permits blastema formation [28]. We followed BM regeneration after re-epithelialization and asked whether it occurred prior to the onset of dermal regeneration in excisional flank wounds. In uninjured skin the BM is visible as a thick fibrous band separating epidermis from dermis and is continuous except where mucous glands interject into the epidermis (Figure 3A). Following re-epithelialization histological staining revealed a thin, immature structure beneath the new epidermis (Figure 3A). The BM continued to mature and was completely regenerated at least 47 days after wounding (Figure 3A; yellow arrows D47). Interestingly, complete regeneration of the BM corresponded to regeneration of the dermis (except for stratum spongiosum) (Figure 3A and Figure 1G).

A) Histological examination of basement membrane (BM) regeneration in axolotls. The uninjured BM is visible as a thick blue-stained fibrous band (yellow arrows). An immature BM has begun to reform (yellow arrow D1) after re-epithelialization and is visible at the wound margin (WM) in contrast to the uninjured BM. The regenerated BM is visible at D47. Yellow arrows at D7 and D21 indicate reforming BM. B) Examination of lamina lucida (laminin) and lamina densa (collagen type IV) during basement membrane regeneration. The uninjured BM is positive for laminin and collagen type IV (yellow arrows) as are the basement membranes surrounding glands and muscle fibers. Following re-epithelialization the basal lamina of the epidermis is negative for laminin and collagen type IV (white arrows) and this is clearly evident at the wound margin (WM). Seven days post injury the BM stains strongly for laminin indicating reformation of the lamina lucida, while staining for collagen type IV is punctuated. The lamina densa is regenerated by D14 based on continuous collagen type IV staining and persists during dermal regeneration.

While adult mammals are incapable of regenerating full thickness skin wounds, fetal mammals exhibit scarless healing of similar type wounds [12]. Similarly, while pre-metamorphic anurans heal scar-free, post-metamorphic anurans have been documented to heal flank wounds through scar formation [29]. Adult axolotls retain several larval skin features (e.g. leydig cells, pseudo-stratified epithelium), thus we asked if these characteristics facilitate their ability to heal wounds scar-free. To test this hypothesis we exploited the fact that normally aquatic axolotls retain the ability to undergo metamorphosis to a terrestrial form through administration of thyroxine and we induced metamorphosis in adult axolotls (controlling for age and size with sibling paedomorphs). Comparing uninjured epidermis between both forms we noted two major differences; first, granular glands that occupied relatively little space in the paedomorph dermis were greatly enlarged and occupied most of the stratum spongiosum while mucous glands appeared similar in form between morphs (Figure 4A and Figure S3A). Second, the epidermis no longer contained leydig cells and had transitioned to a completely stratified epithelium exhibiting a well-defined stratum germinativum, stratum spinosum, stratum granulosum and stratum corneum (Figure 4B).

Complete dermal regeneration was delayed in metamorphs (compare Figure 1G and Figure 4G). While epidermal organs regenerated in both forms after 40 days, the wound bed and underlying muscle still contained densely compacted extracellular matrix in metamorphs (Figure 4G and Figure S3C). After 80+ days the stratum spongiosum had regenerated but the stratum compactum remained incomplete (Figure 4H). After 120 days, the wound site resembled an 80-day regenerating wound in paedomorphs and a few collagen deposits still persisted in the underlying muscle (Figure S3D). Fibrosis was not resolved until at least 148 days and while mucous glands regenerated to pre-wound size, granular glands remained small even after 148 days (Figure S3E). Taken together these findings suggest that flank skin in adult metamorphic axolotls can completely regenerate following FTE wounding, but the time required to regenerate both the stratum compactum and mature granular glands is lengthened compared to paedomorphs.

A-B) Fibronectin (FN) and tenascin-C (TN-C) levels were detected during scar-free healing in paedomorphs and metamorphs using an antibody to axolotl fibronectin and a polyclonal antibody to chick tenascin-C. We detected low levels of FN in the basement membrane at D7, and at the wound margins in both morphs. FN was present during ECM deposition at D14 in the center of the wound bed, but in relatively small amounts. By D21 little FN persisted in the regenerating dermis. B) TN-C was detected at the wound margins, in the basement membrane and surrounding some cells at D7. Fourteen days post injury we detected high levels of TN-C throughout the wound bed and in regenerating muscle. A sharp boundary formed between intact muscle and regenerating muscle. These high levels of TN-C persisted during dermis regeneration. Green fluorescence was used to detect autofluorescing erythrocytes. Epidermis (E), dermis (D), muscle (M), wound margin (WM). 041b061a72


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