Prevalence and Isotypic Complexity of the Anti-Chinese Hamster Ovary Host Cell Protein Antibodies in Normal Human Serum

Host cell-derived protein impurities may be present at low levels in biopharmaceutical products. Antibodies to host cell proteins are present in individuals with no known exposure to these products. In this study, antibodies to drug process-specific Chinese hamster ovary host cell-derived proteins (CHO-HCP) were measured in unexposed individuals using a validated enzyme-linked immunosorbent assay. Samples that tested positive for anti-CHO-HCP reactivity were further characterized to determine the isotypes and IgG subclasses expressed in each positive individual. The specificity of the detected anti-CHO-HCP antibody isotypes was confirmed by the competitive inhibition assay and the uncoated plate specificity testing. These antibody characterization experiments revealed that the prevalent anti-CHO-HCP antibody subclasses were predominantly IgG1 (present in 66.6% of individuals) and IgG2 (60%), with 33.3% positive for IgG3 while IgG4 was detected in low abundance. Forty percent (40%) of the individuals with IgG type reactivity were also identified as IgM-positive. Anti-CHO-HCP-specific IgE was not detected in the assays used in this study. Some individuals exhibited single isotype anti-CHO-HCP reactivity; others contained a combination of multiple antibody isotypes. Data presented in this study demonstrate the isotypic complexity and the high prevalence of anti-CHO-HCP antibodies in human serum with no known exposure to CHO cell-derived therapeutic biologics.     

Sodium channelopathies of skeletal muscle result from gain or loss of function

Five hereditary sodium channelopathies of skeletal muscle have been identified. Prominent symptoms are either myotonia or weakness caused by an increase or decrease of muscle fiber excitability. The voltage-gated sodium channel Na_V1.4, initiator of the muscle action potential, is mutated in all five disorders. Pathogenetically, both loss and gain of function mutations have been described, the latter being the more frequent mechanism and involving not just the ion-conducting pore, but aberrant pores as well. The type of channel malfunction is decisive for therapy which consists either of exerting a direct effect on the sodium channel, i.e., by blocking the pore, or of restoring skeletal muscle membrane potential to reduce the fraction of inactivated channels.     

Verbal-autonomic response dissociations as traits?

Dissociations between subjective and physiological responses to stress are of central interest in coping research. However, little is known about their stability across situations and time. Two experimental sessions - separated by 1 year - were conducted to examine cross-situational consistency and longterm-stability of HR-derived and SCL-derived dissociation scores. In year 1, a speech stressor, the cold pressor and a video stressor (viewing of the speech video) were applied. In year 2, mental arithmetics, anagrams and a torture video were presented. Thirty-five students participated and HR, SCL and negative affect were recorded. For each stressor, standardized changes in negative affect were subtracted from changes in autonomic reactivity (HR and SCL, respectively). Dissociation scores were relatively consistent across the stressors with HR-derived scores exceeding SCL-derived scores. Longterm-stability proved acceptable (r=.61, P<.001 for HR-derived and r=.40, P<.05 for SCL-derived scores). In sum, verbal-autonomic response dissociations show considerable cross-situational and temporal stability and thus might be considered as traits.     

Recurrent high-grade leiomyosarcoma with heterologous osteosarcomatous differentiation

Leiomyosarcomas (LM) of the soft tissue comprise approximately 5–10% of all soft tissue sarcomas. Besides the classic LM, several distinctly uncommon features of the cellular and growth patterns of LM have been described. The term “dedifferentiated LM” has rarely been used in the literature to describe soft tissue LM containing areas of undifferentiated, pleomorphic appearance or detectable heterologous differentiation. We report on a case of high-grade LM with almost entire transition to an osteosarcoma, which was classified as recurrent high-grade LM with heterologous osteosarcomatous differentiation. The identification of areas with osteosarcomatous dedifferentiation in soft tissue sarcomas can be of clinical importance because of a possible change in oncologic treatment strategies.     

Dendritic cell activating peptides induce distinct cytokine profiles

High-mobility group box 1 protein (HMGB1), a DNA-binding nuclear and cytosolic protein, is a pro-inflammatory cytokine released by monocytes and macrophages. HMGB1 as well as its B box domain induce maturation of human dendritic cells (DCs). This report demonstrates that the B box domain induces phenotypic maturation of murine bone marrow-derived dendritic cells (BM-DCs) as evidenced by increased CD86, CD40 and MHC-II expression. The B box domain enhanced secretion of pro-inflammatory cytokines and chemokines: IL-1beta, IL-2, IL-5, IL-8, IL-12 and tumor necrosis factor (TNF)-alpha, but not IL-6 and IL-10. Furthermore, four peptides whose sequences correspond to different regions of HMGB1 induced production of IL-1beta, IL-2 and IL-12 (p70), but not IL-10 and IL-6 in mouse BM-DCs. Interestingly, these peptides differed in their capacity to induce TNF-alpha, IL-5, IL-18 and IL-8. B box domain as well as peptide-activated DCs acted as potent stimulators of allogeneic T cells in a mixed leukocyte reaction. DCs exposed to HMGB1 peptides induced proliferation of ovalbumin-specific syngeneic T cells. These DC-activating peptides could serve as an adjuvant in immunotherapeutic or vaccine context and the selective activity of these different peptides suggests a means to customize the functional properties of DCs.     

Posttraumatic cerebrospinal fluid cyst of the orbit

A 41-year-old woman had a motor vehicle accident and sustained a fracture of the left temporal bone with anterior and middle skull base involvement. After 10 months, she developed persistent right-sided exophthalmus. Orbital computed tomography scans showed a soft tissue mass in the roof of the right orbit with an inferior calcified border. The surgery revealed a cerebrospinal fluid cyst with intracranial communication through the fistula in the posterior orbital roof.     

Lymphocytes are detrimental during the early innate immune response against Listeria monocytogenes

Mice deficient in lymphocytes are more resistant than normal mice to Listeria monocytogenes infection during the early innate immune response. This paradox remains unresolved: lymphocytes are required for sterilizing immunity, but their presence during the early stage of the infection is not an asset and may even be detrimental. We found that lymphocyte-deficient mice, which showed limited apoptosis in infected organs, were resistant during the first four days of infection but became susceptible when engrafted with lymphocytes. Engraftment with lymphocytes from type I interferon receptor-deficient (IFN-alphabetaR(-/-)) mice, which had reduced apoptosis, did not confer increased susceptibility to infection, even when the phagocytes were IFN-alphabetaR(+/+). The attenuation of innate immunity was due, in part, to the production of the antiinflammatory cytokine interleukin 10 by phagocytic cells after the apoptotic phase of the infection. Thus, immunodeficient mice were more resistant relative to normal mice because the latter went through a stage of lymphocyte apoptosis that was detrimental to the innate immune response. This is an example of a bacterial pathogen creating a cascade of events that leads to a permissive infective niche early during infection.     

CD40 induces macrophage anti-Toxoplasma gondii activity by triggering autophagy-dependent fusion of pathogen-containing vacuoles and lysosomes

Many intracellular pathogens, including Toxoplasma gondii, survive within macrophages by residing in vacuoles that avoid fusion with lysosomes. It is important to determine whether cell-mediated immunity can trigger macrophage antimicrobial activity by rerouting these vacuoles to lysosomes. We report that CD40 stimulation of human and mouse macrophages infected with T. gondii resulted in fusion of parasitophorous vacuoles and late endosomes/lysosomes. Vacuole/lysosome fusion took place even when CD40 was ligated after the formation of parasitophorous vacuoles. Genetic and pharmacological approaches that impaired phosphoinositide-3-class 3 (PIK3C3), Rab7, vacuolar ATPase, and lysosomal enzymes revealed that vacuole/lysosome fusion mediated antimicrobial activity induced by CD40. Ligation of CD40 caused colocalization of parasitophorous vacuoles and LC3, a marker of autophagy, which is a process that controls lysosomal degradation. Vacuole/lysosome fusion and antimicrobial activity were shown to be dependent on autophagy. Thus, cell-mediated immunity through CD40 stimulation can reroute an intracellular pathogen to the lysosomal compartment, resulting in macrophage antimicrobial activity.