Microbes in Skin Probiotics

Introduction

The microbiome of the human body is incredibly diverse, and plays a key role in affecting the health of the individual. The skin microbiota is the second largest microbiome on the human body, after the gut, and is home to a vast array of bacteria, fungi, and viruses (Byrd et al, 2018). In recent years more and more research has been done to understand how the skin’s microbiome affects human wellbeing, and how to use this knowledge to create better probiotic products that can treat different skin conditions.

In a healthy skin microbiota, it is found that the resident communities on the skin remained largely stable over the course of multiple years (Byrd et al, 2018). Many skin diseases are associated with changes in the microbiota, where there is a loss in diversity or decrease in stability, although whether these changes play a causative role in development of skin conditions is a question still under investigation. Skin probiotics help by administering living or dead microorganisms into the hosts’ microbiome to combat these issues and strengthen innate immunity. Most approaches to treat skin conditions administer probiotics orally, but some involve direct application of probiotic microorganisms or microbial products directly to the skin. Conditions such as acne, skin hypersensitivity, eczema, atopic dermatitis, and even skin aging and cancer all have the potential for treatment or prevention by probiotics (Roudsari et al, 2015).

Review of Research on Skin Probiotic Microorganisms

Nitrosomonas eutropha

Both oral and topical probiotics have become commonly used in skincare to treat skin conditions or signs of aging. Ammonia-oxidizing bacteria (AOB), like Nitrosomonas eutropha, have been getting a lot of attention, due to healing properties associated with nitrogen compounds. Nitrosomonas eutropha has been a popular addition to acne, eczema and anti-aging products. This bacteria oxidizes ammonia from sweat, thus producing nitrite and nitric oxide, which have been associated with increased vasodilation, anti-inflammatory, and wound-healing properties. In previous preclinical studies, N.eutropha improved acne by outpopulating Propionibacterium acnes and visibly improving healing/cicatrization. Some studies in course aim to make N.eutropha aerosols for mild-moderate acne.

In a separate study conducted with aerosolized live N. eutropha on wrinkles and pigmentation, significant results were obtained after treating 29 participants (ages 23-41) with high and low concentration N.eutropha. To create a baseline, all participants had clear skin. Participants were excluded if they had been on antibiotics (topical or systemic) recently, isotretinoins, retinoids or any active skin treatment. Pregnant and participants switching brands of oral contraceptives were also excluded. Treatment with N.eutropha had significantly improved wrinkle severity and depth on the glabella, forehead and overall face. Improved pigmentation in the forehead and glabella, not so much in the overall face were also observed. Overall, the high concentration treatment had improved skin condition significantly more than the low concentration (Notay et al., 2019).

Modern hygienic lifestyles have driven atopic diseases due to lack of immune maturation. Exposure to microbes is needed for proper/robust maturation of the immune system, and lack of thereof has been highly associated with atopic diseases. A common feature of atopic diseases is the exacerbated, systemic type 2 inflammatory reaction inducing high amounts of immunoglobulin E (IgE) by B cells, eosinophilia, and many other immune agents. Therapies that target Th2 cells and associated cytokines (IL-5, IL-13, IL-4) have shown to be effective against this pathway (Maura et al., 2021). To target the pathway, the focus was shifted to bacteria that have the ability to modulate the Th2 pathway, like the popular ammonia-oxidizing bacteria (AOB). Often found in farm soil, due to animal waste and high concentrations of fertilizers, AOB have been depleted in human microbiomes over the years. A study on the subject found that N.eutropha inhibits the polarization of CD4+ T to Th2 in human primary immune cells, and does not require the Th1 pathway to achieve this goal (Maura et al., 2021). This study was one of the first ones to support the use of AOB in the skin, for the treatment of Th2-mediated atopic diseases.

N.eutropha has also been tested against keratosis pilaris, during a focused clinical trial on cutaneous microflora. The objective/goal was to restore dermal microflora by colonizing the skin with a purified strain of N.eutropha, an evolutionary important source of Nitric oxide (anti -inflammatory and antimicrobial) in the body (Ny et al., 2018). Satisfactory results would give new direction to the quest for better skin health.

Bifidobacterium lactis

Bifidobacterium lactis is a gram-positive rod-shaped bacterium found in the large intestines of cows, cats, and humans. This bacteria was isolated from a breastfed infant in the 1980s. It is a subspecies deriving from Bifidobacterium animalis. This bacterium was sequenced because of the boom of probiotic use and its possibility to be a good bacteria to add to the human gut microbiome. B. lactis is considered a good probiotic because it expresses fermentation activity, it has a high aero tolerance and good stability. It can be found in formula, supplements, and fermented milk products globally. It also can go undetected orally because it does not have a taste or a specific texture that would make it a difficult consumable. It is able to survive the stressful environment of the gastrointestinal tract by having a high acid and bile salt tolerance. A specific strain of B. lactis named BB-12 expresses all of the main qualities of being a productive probiotic. It can inhibit pathogens, enhance barrier function, and aid in immune interactions (Jungerson et. al 2014).

Probiotics are able to interact with the immune system due to the interactions through the colonization of the gut mucosa and the gut-associated lymphatic tissue. Keeping the gut in a proper symbiotic environment reduces the event of gut dysbiosis that can lead to inflammatory events in other parts of the body including the GI tract. The metabolic processes of B. lactis can be attributed to the release of short-chain fatty acids (SCFA). Since B. lactis is a fermenter it can produce SCFAs as a byproduct of fiber breakdown. These SCFAs are used by the body and can decrease skin inflammation and other inflammatory processes due to their ability to influence macrophages, neutrophils, and other leukocytes. The Bifidobacteria family has also been known to help encourage the growth of immune cells like dendritic cells and T cells and are introduced early in life through breastfeeding. B. lactis and other organisms in this family have been able to decrease some inflammatory skin conditions as well as food allergies (Mann et al. 2020).

In a study to look at the use of probiotics to treat atopic dermatitis, commonly known as eczema, researchers used B. lactis in association with a Lactobacillus strain. Their hypothesis was that a probiotic regimen of B. lactis and Lactobacillus casei would cause improvement in atopic dermatitis symptoms. A population of patients ranging from 4-17 years of age took the placebo or probiotic for 12 weeks. Following the 12-weeks the patients that received the probiotic had an increase in B. lactis found in their stool samples. When looking at this increase it lead to the belief that B. lactis has the ability to decrease the gut bacteria that typically will aggravate atopic dermatitis. The decrease in the bacteria aggravating AD and the increase in the B. lactis lead to a lower percentage of AD symptoms in these patients that received the probiotic course of treatment (Climent et. al 2021). This is a newer study and in the past opposing results have been found. Another study aimed at examining AD in infants, found that Lactobacillus decreased AD flare-ups, but B. lactis had no overall effect. If there was an effect found it would not have been beneficial due to B. lactis being absent in stool samples when they were collected more than 3 days after probiotic ingestion (Wickens et al 2008).

In conclusion, a growing body of research shows evidence that B. lactis helps to cultivate a healthy gut and stimulates down-regulation of inflammatory processes that are unnecessary in various systems in the body. It is able to establish a balanced gut microbiome and is a beneficial probiotic in some studies looking at atopic dermatitis. Due to its capability to communicate with the immune system and produce SCFAs it does serve as an aid to decrease inflammation which may translate to skin inflammation in the future.

Lactobacillus rhamnosus

Lactobacillus rhamnosus is a bacteria that is naturally found in our gut. It is an anaerobic or facultative anaerobic Gram-positive rod that is commonly found in the human gastrointestinal tract and vaginal tract (Albarillo, F. S et al., 2020). L. rhamnosus is being researched as a probiotic-based treatment for atopic dermatitis. Atopic dermatitis is a chronic inflammatory skin disease that is due to mutations of the filaggrin gene, which is located in the epidermal differentiation complex. The skin inflammation can be caused by many factors such as allergens, dust mites, changes in weather, and stress. This skin condition affects 5-20% of children worldwide and the incidence continues to increase each year. (Wu, Y.-J et al., 2017). These have been used in medical practice as immunomodulators. Lysates of L. rhamnosus can increase tight-junction barrier resistance by modulating the specific protein complexes. Studies have been conducted and shown that Lactobacilli lysates can re-epithelialize keratinocytes (Puebla-Barragan, S., & Reid, G. 2021) A study was done to test the lysates of L. rhamnosus on skin barrier function in reconstructed human epidermis, Keraskin. Keraskin was purchased from Biosolution Co. and consists of human keratinocytes as a functional multilayered epidermis with the essential lipid composition. A L. rhamnosus (LR) lysate when used topically in this study has been shown to increase epidermal differentiation markers. The LR lysate, when applied to Keraskin every other day for 16 days, showed that the treated tissues have more ordered and denser stratum corneum when compared with the control. The treated tissues also underwent immunofluorescence staining with the antibodies against two specific tight junction proteins: claudin1 and occludin. The IF images showed an increase in both tight junction molecules after treatment with the LR lysate. To further analyze this, immunohistochemistry with antibodies against cytokeratin 5, 1, 10, loricrin, and filaggrin was performed. K5 and K1 showed an advancement into the upper layer of the epidermis along with increasing intensity of filaggrin in LR treated Keraskin (Jung, Y.-O et al., 2019).

A randomized double-blind, placebo-controlled study was conducted at two sites in Taiwan to investigate the efficacy of L. rhamnosus in children from the ages 4-48 months. The study suggests a positive effect of probiotic supplementation in patients with atopic dermatitis (Wu, Y.-J et al., 2017). The data showed a greater decrease in the intensity and severity of AD for the population who received at least one dose of the treatment of Lactobacillus rhamnosus. However, there were many limitations because the onset of probiotics may be more significant if the treatment lasts more than 8 weeks. Also, AD can have recurrence and fluctuates in the short term. However, when treatment with L. rhamnosus was used orally in combination with topical corticosteroids, it showed a synergistic effect in this study (Wu, Y.-J et al., 2017).

Another study was performed in Wellington and Auckland, New Zealand to test that probiotic supplementation by L. rhamnosus administered to mothers from 14 to 16 weeks gestation until delivery and continuing until 6 months postpartum, if breastfeeding, will reduce the prevalence of infant eczema and atopic sensitization and the prevalence of maternal GDM, BV, and GBS vaginal colonization before birth (Barthow, C et al., 2016). However this study has not provided its data as it is ongoing.

Overall, L. rhamnosus is currently being researched for usage as a probiotic to help treat skin conditions like atopic dermatitis. Since this bacteria is naturally found in the gastrointestinal tract as well as the vaginal tract, it shows promise as a potential treatment for this condition. Studies have been done like the ones above that have strong data supporting the use of L. rhamnosus, but more studies should be completed in order to show replicable data in human based models.

Lactobacillus acidophilus

Lactobacillus acidophilus is a gram positive rod-shaped bacteria that is typically found in human and animal gastrointestinal tract and mouth (Badet and Thibauld, 2008). While more well known for its probiotic uses in dairy products such as yogurt, L. acidophilus has been recently receiving attention for its uses in skin care as an anti-wrinkle agent.

L. acidophilus was first noticed for potential skin treatments alongside the proposed gut-brain-skin theory by John H. Stokes and Donald M. Pillsbury around the 1930s. The theory proposed that gastrointestinal microbiome and function were tied to mental health disorders and skin conditions such as acne. Chronic emotional states alter the gastrointestinal tract, which disturb the intestinal microbial flora and increase the risk for intestinal permeability. The resulting impact is an increase in inflammation to the skin. The proposed solution was 'the direct introduction of acidophil organisms in cultures,' which could be distributed with tablets or drinks. Formal studies were conducted starting in the 1960s, often with oral probiotic tablets containing L. acidophilus in combination with other bacteria such as L. bulgaricus and B. bifidum. While flawed, these studies indicated a strong correlation between the intake of bacteria and the reduction of inflammation acne and other skin lesions (Bowe WP et. al. 2011).

More recent studies have turned their focus on tyndalized L. acidophilus's effects on UV damaged skin as an anti-wrinkle agent. A study in 2016 focused on UVB radiation on the skin of hairless mice and the potential effects from orally administered tyndalized L. acidophilus tablets. Tyndallization is an outdated sterilization process that heats a solution at or just below boiling temperatures, and is used today to kill bacteria as well as any endospores possibly contained inside. The mice exposed to UVB rays experienced significant skin dehydration and transepidermal water loss, however thanks to frequent ingesting L. acidophilus beforehand, some of the hydration and water retention was retained in the skin and the overall damage was not as severe. Skin samples also revealed a more even collagen distribution among mice that received the L. acidophilus tablets, resulting in a lower level of wrinkling as a result of UV irradiation. A western blot has revealed that the MMPs, specifically MMP-1 and MMP-9, alongside signaling parts involved in the p38 MAPK pathway, have a reduced expression. These enzymes are strongly expressed as a result of UV irradiation, so further investigation on how L. acidophilus reduces the expression could be explored (Im AR et al. 2016).

Further studies have explored L. acidophilus effects on human cells alongside mice cells. Human keratinocytes and human dermal fibroblast cells were exposed to UVB irradiation to observe the skin's biological response to tyndallized L. acidophilus, specifically L. acidophilus-KCCM12625P. The cell viability of HaCaT and HDF cells saw a recovery from ROS generation due to UVB irradiation after exposure to L. acidophilus, indicating antioxidant effects. ROS induced wrinkles as a result of induction of MMPs and elastase enzymes were also inhibited in a dose-dependent manner. The melanogenesis of B16F10 mice cells through UVB irradiation was also observed, and indicated that L. acidophilus does not directly affect tyrosinase activity but has anti-melanogenesis effects through regulation of cAMP signaling pathways (Lim HY et. al. 2020).

The overarching conclusion is that L. acidophilus does have potential positive benefits to skin care, with a particular emphasis on skin protection from UVB irradiation damages. L. acidophilus has been found to aid in the reduction of dehydration, water retention, and inhibiting ROS induced wrinkles from mice models. This is achieved through impacting the p38 MAPK and cAMP pathways. Human cell culture models also saw a recovery from ROS induced wrinkles through the known molecular "aging" markers. However, studies using human subjects are either lacking or flawed. The early studies often either did not have a control for comparison, or were difficult to evaluate due to different language or accessibility. Nevertheless, these findings provide a solid basis for future analysis on L. acidophilus's role in skin care.

Lactobacillus paracasei

Acne is an inflammatory disorder involving the pilosebaceous unit (Porubsky et al, 2018). The pathophysiology of acne is mediated by excess sebum production, follicular hyperkeratinization, hyper-colonization with Cutibacterium acnes, and inflammation (Nole et al, 2014). These factors can be aggravated by stress, which leads to alteration of the intestinal lining by encouraging dysbiosis. Fifty four percent of patients with acne vulgaris have marked alterations within intestinal flora (Nole et al, 2014). Disruption of gut flora can lead to conditions including constipation, which greatly reduces the amount of Lactobacilli and Bifidobacteria within the intestinal microbiome. Some benefits of probiotics are reducing inflammation, and reduction in sebum production which inhibits hyper colonization of P. acnes. (Nole et al, 2014) Lactobacillus paracasei is a Gram positive, facultative anaerobe, non-spore forming bacteria. They form a part of the normal human microbiota of the oral, gastrointestinal, and female genital tracts (Tang et al, 2021). Several studies have investigated the potential to use L. paracasei as a probiotic agent for the treatment of acne. Probiotics have been used to provide benefits in reducing acne, and previous studies have shown that L. paracasei can reduce inflammation. L. paracasei supernatant was shown to have an inhibitory effect on antibiotic resistant and non-resistant Cutibacterium acnes, which is a bacteria that lives on the human skin, mostly within hair follicles on the face and neck. In a human study, it was found that when applied topically, L. paracasei supernatant reduced the symptoms of mild-to-moderate acne vulgaris, comparable to other known lotions to help with acne (Sathikulpakdee et al, 2022). Additional work has shown a decrease in skin inflammation following treatment with L. paracasei probiotics, with alterations in edema, vasodilation, mast cell degranulation, and TNF-alpha release induced by P. acnes(BPharm and Suyenaga, 2018). The effects of the skin barrier reconstruction and inflammatory inhibition do not only have beneficial effects for acne, but also helps with Atopic Dermatitis, which is correlated with cutaneous inflammation and a deficient skin barrier (BPharm and Suyenaga, 2018). Dysbiosis of the gastrointestinal tract can lead to multiple different health problems throughout the body. The gastrointestinal tract houses the largest population of commensal bacteria, and within this reservoir of this bacteria lies the central theme to the gut-brain-skin axis. Hypochloridia and small intestinal bacterial overgrowth (SIBO) are two conditions that demonstrate an association with cutaneous pathology and mental health. In a previous study, probiotic use of Lactobacillus paracasei successfully reduced SIBO, which showed improvement of both GI health and cutaneous symptoms. (Porubsky et al, 2018) In summary, the work described here suggests that when taken orally or topically, Lactobacillus paracasei can help combat the inflammation that contributes to acne. It has the ability to inhibit the growth of Cutibacterium acnes when applied topically and when taken orally, was shown to have multiple benefits including reduction of inflammation, which is a driving factor in acne development. It was also shown to reduce small intestinal bacterial overgrowth, which led to improvement of cutaneous symptoms. By introducing probiotics containing Lactobacillus paracasei it can be seen that there is a reduction of acne and overgrowth of the bacteria that causes acne.