Large scale expansion of gamma 9 delta 2 T lymphocytes: Innacell gamma delta cell therapy product.

gamma9delta2 T lymphocytes are non-conventional lymphocytes presenting a direct cytotoxic effect against a broad range of tumour targets. These cells also secrete inflammatory cytokines that can boost the other components of the immune system. In contrast to conventional CD8(+) T cells, the cytotoxic effect of gamma9delta2 T lymphocytes does not depend on the expression of major histocompatibility complex molecules by target tumour cells. INNACELL gammadeltatrade mark is a cell therapy product obtained by ex vivo amplification of mononuclear cells. The stimulation is achieved by a specific synthetic agonist of gamma9delta2 T lymphocytes, bromohydrin pyrophosphate (BrHPP). After a single stimulation with BrHPP, gamma9delta2 T lymphocytes are expanded for 2 weeks in a closed system in culture medium with interleukin-2 (IL-2). On day 15, cells are washed and harvested in 4% human serum albumin. In this manufacturing process, the total cell population is expanded by approximately 10-fold and gamma9delta2 T lymphocytes undergo a specific 1000-fold expansion, corresponding to a gamma9delta2 T lymphocyte enrichment of more than 70% at the end of the culture. This manufacturing process is much simpler than most current cellular therapy approaches using conventional CD8(+) T-cell lines or clones: there is no final or initial separation, no purification step and no use of feeder cells; the specific T-cell receptor-mediated signal provided by BrHPP is sufficient to trigger the IL-2-dependent expansion of the gamma9delta2 subset, which then becomes predominant in the cell culture in large amounts.

In vitro modulation of TLR-2, CD1d and IL-10 by adapalene on normal human skin and acne inflammatory lesions.

The anti-inflammatory mechanisms of adapalene, a synthetic retinoid used for the treatment of acne patients, are partially understood. They seem particularly related to the modulation of the non-specific immunity. Recent studies have shown that Toll-like receptor (TLR)-2 expression, a receptor of the innate immune system, was increased in acne lesions and could play an essential role in acne-linked inflammation. The aim of our study was to investigate the new mechanisms of the anti-inflammatory activity of adapalene in vitro, and more specifically the modulatory effect of adapalene on the expression of TLR-2, CD1d, a cell surface glycoprotein that plays a role as antigen-presenting molecules and is responsible for the development of cutaneous inflammation, and also on the expression and the secretion of the anti-inflammatory interleukin (IL)-10 cytokine. Both explants of normal human skin and explants of acne patients were incubated with adapalene (10(-7) or 10(-6) M) or the control medium for 24 h. Evaluation of epidermal expression by immunohistochemistry showed a decreased expression of TLR-2 and IL-10 in explants of normal skin and explants of acne with adapalene. On the contrary, adapalene increased CD1d expression in explants of acne patients. Thus, adapalene can modulate the epidermal immune system by increasing the CD1d expression and by decreasing the IL-10 expression by keratinocytes. Moreover, these modulations could increase the interactions between dendritic cells and T lymphocytes and could strengthen the antimicrobial activity against Propionibacterium acnes. The decreased expression of TLR-2 by the keratinocytes can contribute to explain the anti-inflammatory activity of adapalene observed in clinical practice.

Modulation in vitro of keratinocyte integrins by interferon-alpha and interferon-gamma.

BACKGROUND: Interferon-alpha and -gamma are glycoproteins with antiviral and immunoregulatory properties. In vitro studies have shown a role for these cytokines in the regulation of epidermal keratinocyte growth and differentiation. In the same way, integrins are adhesion molecules which regulate keratinocyte proliferation and differentiation. AIM: To determine whether the regulatory activity of interferons on keratinocyte proliferation and differentiation is related to a modulation of keratinocyte integrins. METHODS: Two different methods were used: monolayers and reconstituted skin, incubated either with 1,200 U/mL interferon-alpha or 500 U/mL interferon-gamma or control medium for 48 h. The integrin expression was assessed by flow cytometry and immunohistochemistry. RESULTS: In monolayers, only the alpha3 subunit was significantly inhibited by interferon-gamma. In reconstituted skin, where keratinocytes are differentiated, both interferons had an inductive effect on beta1 expression and interferon-alpha had an inhibitory effect on alpha6 expression. CONCLUSION: Interferon-alpha and -gamma induce a modulatory effect on alpha3, alpha6 and beta1 which appears to be related to the state of differentiation. Moreover, the decreased expression of alpha6 and alpha3 could be one of the mechanisms involved in the formation of bullous lesions during long-term interferon therapy.