Several groups are investigating the mechanisms of action of therapeutic immunoglobulins (IVIg) in order to improve their use. In vitro models such as CD40-CD154 interaction are necessary to study the physiological re...Several groups are investigating the mechanisms of action of therapeutic immunoglobulins (IVIg) in order to improve their use. In vitro models such as CD40-CD154 interaction are necessary to study the physiological response of human B lymphocytes to IVIg. Human B lymphocytes treated with IVIg triggers a rapid phospho-rylation (<1 h) of extracellular-regulated kinases 1 and 2 (ERK1/2), which subsequently results in increased differentiation and decreased pro-liferation. However, the modulation of human lymphocyte physiology by IVIg is a gradual and cumulative process and requires long-term experimentation. Differentiation of human B lymphocytes into Ig-secreting cells can be evaluated both at the transcription and translation levels. The secretion of immunoglobulins can be assessed using ELISA or ELISPOTS whereas expression of immunoglobulin genes can be measured using semi-quantitative or quantitative PCR methods. We hereby report a comparison of these methods to explain how contradictory observations towards IVIg effects could result from their use. Our results indicate that ELISA and ELISPOTS will provide consistent observations by opposition to real-time PCR quantification. Besides, the reliability of each of these me-thods remained dependent on the stimulation period as well as the preparation of cellular extracts or cell samples following IVIg-treatment.展开更多
Sympathetic neuronal differentiation is associated with favorable prognosis of neuroblastoma （NB）, the most common extra-cranial solid tumor of early childhood. Differentiation agents have proved useful in clinical ...Sympathetic neuronal differentiation is associated with favorable prognosis of neuroblastoma （NB）, the most common extra-cranial solid tumor of early childhood. Differentiation agents have proved useful in clinical protocols of NB treatment, but using them as a sole treatment is not sufficient to induce tumor elimination in patients. Therefore, complementary approaches, such as immunotherapy, are warranted. Here we demonstrate that differentiation of NB cell lines and ex vivo isolated tumor cells in response to physiological or pharmacological stimuli is associated with acquisition of increased antigenicity. This manifests as increased expression of surface major histocompatibility class I complexes and ICAM-1 molecules and translates into increased sensitivity of NB cells to lysis by cytotoxic T lymphocytes （CTLs） and natural killer （NK） cells. The latter is paralleled by enhanced ability of differentiated cells to form immune conjugates and bind increased amounts of granzyme B to the cell surface. We demonstrate, for the first time, that, regardless of the stimulus applied, the differentiation state in NBs is associated with increased tumor antigenicity that enables more efficient elimination of tumor cells by cytotoxic lymphocytes and paves the way for combined application of differentiation-inducing agents and immunotherapy as an auxiliary approach in NB patients.展开更多
文摘Several groups are investigating the mechanisms of action of therapeutic immunoglobulins (IVIg) in order to improve their use. In vitro models such as CD40-CD154 interaction are necessary to study the physiological response of human B lymphocytes to IVIg. Human B lymphocytes treated with IVIg triggers a rapid phospho-rylation (<1 h) of extracellular-regulated kinases 1 and 2 (ERK1/2), which subsequently results in increased differentiation and decreased pro-liferation. However, the modulation of human lymphocyte physiology by IVIg is a gradual and cumulative process and requires long-term experimentation. Differentiation of human B lymphocytes into Ig-secreting cells can be evaluated both at the transcription and translation levels. The secretion of immunoglobulins can be assessed using ELISA or ELISPOTS whereas expression of immunoglobulin genes can be measured using semi-quantitative or quantitative PCR methods. We hereby report a comparison of these methods to explain how contradictory observations towards IVIg effects could result from their use. Our results indicate that ELISA and ELISPOTS will provide consistent observations by opposition to real-time PCR quantification. Besides, the reliability of each of these me-thods remained dependent on the stimulation period as well as the preparation of cellular extracts or cell samples following IVIg-treatment.
文摘Sympathetic neuronal differentiation is associated with favorable prognosis of neuroblastoma （NB）, the most common extra-cranial solid tumor of early childhood. Differentiation agents have proved useful in clinical protocols of NB treatment, but using them as a sole treatment is not sufficient to induce tumor elimination in patients. Therefore, complementary approaches, such as immunotherapy, are warranted. Here we demonstrate that differentiation of NB cell lines and ex vivo isolated tumor cells in response to physiological or pharmacological stimuli is associated with acquisition of increased antigenicity. This manifests as increased expression of surface major histocompatibility class I complexes and ICAM-1 molecules and translates into increased sensitivity of NB cells to lysis by cytotoxic T lymphocytes （CTLs） and natural killer （NK） cells. The latter is paralleled by enhanced ability of differentiated cells to form immune conjugates and bind increased amounts of granzyme B to the cell surface. We demonstrate, for the first time, that, regardless of the stimulus applied, the differentiation state in NBs is associated with increased tumor antigenicity that enables more efficient elimination of tumor cells by cytotoxic lymphocytes and paves the way for combined application of differentiation-inducing agents and immunotherapy as an auxiliary approach in NB patients.
