Journal: Alcohol (Fayetteville, N.Y.)

Article Title: The Impact of Alcohol Use Disorders on Pulmonary Immune Cell Inflammatory Responses to Streptococcus pneumoniae

doi: 10.1016/j.alcohol.2018.08.016

Figure Lengend Snippet: Freshly collected alveolar macrophages (AMs) from non-AUD subjects (n=14) and AUD subjects (n=17) were cultured ex vivo with 5 μg heat-killed S. pneumoniae (Sp); cell culture supernatants were collected after two, 18, and 24 hours had elapsed. Interferon (IFN)-γ, interleukin (IL)-1β, IL-6, and tumor necrosis factor (TNF)-α were measured in cell culture supernatants. (2A) AUD subjects exhibited higher secretion of IFNγ, but this did not achieve statistical significance (p=0.10). IFNγ quantity increased significantly over time for both subject types (p<0.0001). (2B) No significant differences in secretion of IL-1β were observed between different subject types, however, secretion of IL-1β increased over time (p=0.0003). (2C) Secretion of IL-6 by AMs from subjects with AUDs tended to be higher (p=0.12). Secretion of IL-6 increased significantly over time in each subject group (p<0.0001). (2D) Secretion of TNFα did not differ between groups (p=0.26), but did increase in each group of subjects over time (p<0.0001).

Article Snippet: In some experiments, heat-killed Streptococcus pneumoniae (Strain JY2008, ATCC, Manassas, VA) was used in specific quantities over delineated time points.

Techniques: Cell Culture, Ex Vivo

Journal: Alcohol (Fayetteville, N.Y.)

Article Title: The Impact of Alcohol Use Disorders on Pulmonary Immune Cell Inflammatory Responses to Streptococcus pneumoniae

doi: 10.1016/j.alcohol.2018.08.016

Figure Lengend Snippet: Freshly collected alveolar macrophages (AMs) from non-AUD subjects (n=14) and AUD subjects (n=17) were cultured in the presence of heat-killed S. pneumoniae protein, at doses ranging from 0 μg to 10 μg. Cell culture supernatants were collected at 18 hours. Interferon (IFN)-γ, interleukin (IL)-1β, IL-6, and tumor necrosis factor (TNF)-α were measured in cell culture supernatants. (3A) IFNγ secretion by AMs rose with exposure to increasing doses of pneumococcal protein (p=0.0002). IFNγ secretion was more elevated in the AUD group (p=0.008). (3B) IL-1β secretion by AMs rose with increasing doses of pneumococcal protein (p=0.02), but was not different between non-AUD and AUD groups. (3C) IL-6 secretion rose with increasing pneumococcal protein doses (p<0.0001), but was not different between non-AUD and AUD groups. (3D) TNFα secretion rose with increasing pneumococcal protein doses (p<0.0001), but was not different between non-AUD and AUD groups.

Article Snippet: In some experiments, heat-killed Streptococcus pneumoniae (Strain JY2008, ATCC, Manassas, VA) was used in specific quantities over delineated time points.

Techniques: Cell Culture

Journal: Alcohol (Fayetteville, N.Y.)

Article Title: The Impact of Alcohol Use Disorders on Pulmonary Immune Cell Inflammatory Responses to Streptococcus pneumoniae

doi: 10.1016/j.alcohol.2018.08.016

Figure Lengend Snippet: Freshly collected alveolar macrophages (AMs) from non-AUD subjects (n=15) and AUD subjects (n=15) were cultured up to 42 hours, with and without the addition of heat-killed S. pneumoniae (Sp, 10μg). In a subset of wells, N-acetylcysteine (NAC) was added after 18 hours of exposure to S. pneumoniae. Cell culture supernatants were collected at the 18 hour and 42 hour time points for analysis of interferon (IFN)-γ, interleukin (IL)-1β, IL-6, and tumor necrosis factor (TNF)-α. (4A) At the 18 hour time point, unstimulated AUD subjects’ AMs secreted more IFNγ (p=0.02) (denoted with asterisk). With pneumococcal protein stimulation, IFNγ secretion by AMs rose significantly between the 18 and 42 hour time points (p=0.002). The addition of NAC at 18 hours during the 42-hour time course was associated with less IFNγ in cell culture supernatants compared to non-NAC treated AMs at 42 hours (p<0.0001 for both non-AUD and AUD subjects). (4B) After pneumococcal protein stimulation, IL-1β secretion by AMs rose significantly between the 18 and 42 hour time points (p=0.0004). Compared to cell culture supernatants at 42 hours without NAC, the addition of NAC at 18 hours during the 42-hour time course was associated with a non-significant rise in supernatant IL-1β among non-AUD subjects (p=0.06), and significantly higher IL-1β in AUD subjects (p=0.009). Cell culture supernatants from pneumococcal protein-stimulated, NAC-treated AMs at 42 hours compared supernatants from pneumococcal protein-stimulated AMs at 18 hours contained IL-1β values that were higher both in non-AUD (p=0.003) and AUD subjects (p=0.01). (4C) At the 18 hour time point, unstimulated AUD subjects’ AMs secreted more IL-6 (p=0.03) (denoted with asterisk). With pneumococcal protein stimulation, IL-6 secretion by AMs rose significantly between the 18 and 42 hour time points (p=0.009). The addition of NAC at 18 hours during the 42-hour time course was associated with substantially less IL-6 secretion than that by non-NAC treated AMs at 42 hours (p=0.0001 for both non-AUD and AUD subjects). Supernatant IL-6 quantities at the 18 hour time point were significantly higher than those in supernatants from pneumococcal protein-stimulated, NAC-treated AMs in both subject types (p=0.05 for non-AUD, p=0.0001 for AUD). (4D) With pneumococcal protein stimulation, TNFα secretion by AMs rose significantly between the 18 and 42 hour time points (p=0.004). The addition of NAC at 18 hours during the 42-hour time course was associated with substantially less TNFα in supernatants than what was measured in non-NAC treated AM culture supernatants at 42 hours (p=0.0001 for both non-AUD and AUD subjects), with values that were significantly less than AM culture supernatants at the 18 hour time point (p=0.02 for non-AUD, and p=0.0006 for AUD). * indicates p≤0.03 between non-AUD and AUD subjects, without pneumococcal stimulation, at 18 hours in culture. # indicates p<0.0001 between NAC and non-NAC treated conditions at 42 hours. & indicates p≤0.05 between non-NAC-treated 18-hour condition and NAC-treated, 42-hour condition.

Article Snippet: In some experiments, heat-killed Streptococcus pneumoniae (Strain JY2008, ATCC, Manassas, VA) was used in specific quantities over delineated time points.

Techniques: Cell Culture

Journal: Alcohol (Fayetteville, N.Y.)

Article Title: The Impact of Alcohol Use Disorders on Pulmonary Immune Cell Inflammatory Responses to Streptococcus pneumoniae

doi: 10.1016/j.alcohol.2018.08.016

Figure Lengend Snippet: Peripheral blood mononuclear cells (PBMCs) from non-AUD subjects (n=8) and AUD subjects (n=8) were cultured in the presence of 5 μg heat-killed S. pneumoniae (Sp); cell culture supernatants were collected at two, 18, and 24 hours. Interferon (IFN)-γ, interleukin (IL)-1β, IL-6, and tumor necrosis factor (TNF)-α were measured in cell culture supernatants. (5A) For IFNγ, AUD subjects exhibited non-significantly higher secretion (p=0.10). Values increased significantly over time (p<0.0001). (5B) For IL-1β, no significant between-groups differences in secretion were observed (p=0.97), however, secretion increased over time (p=0.002). (5C) For IL-6, secretion values from AUD PBMCs did not differ between groups (p=0.35), but secretion increased over time (p=0.003). (5D) For TNFα, values did not differ between groups (p=0.54), and but did increase in subjects over time (p=0.007).

Article Snippet: In some experiments, heat-killed Streptococcus pneumoniae (Strain JY2008, ATCC, Manassas, VA) was used in specific quantities over delineated time points.

Techniques: Cell Culture

Journal: Alcohol (Fayetteville, N.Y.)

Article Title: The Impact of Alcohol Use Disorders on Pulmonary Immune Cell Inflammatory Responses to Streptococcus pneumoniae

doi: 10.1016/j.alcohol.2018.08.016

Figure Lengend Snippet: Peripheral blood mononuclear cells (PBMCs) from non-AUD subjects (n=8) and AUD subjects (n=8) were cultured in the presence of heat-killed S. pneumoniae (Sp), at doses ranging from 0 μg to 10 μg. Cell culture supernatants were collected at 18 hours, and interferon (IFN)-γ, interleukin (IL)-1β, IL-6, and tumor necrosis factor (TNF)-α were measured. (6A) IFNγ secretion over the dose range increased with increasing doses of pneumococcal protein (p=0.04), but values between groups did not differ. (6B) IL-1β secretion increased with increasing pneumococcal protein doses (p=0.004), but were not different between non-AUD and AUD groups. (6C). IL-6 secretion increased with increasing pneumococcal protein doses (p=0.004), but were not different between non-AUD and AUD groups, except at the 0μg (media only) condition (p=0.04, asterisk). (6D) TNFα secretion increased with increasing pneumococcal protein doses (p=0.001), but were not different between non-AUD and AUD groups.

Article Snippet: In some experiments, heat-killed Streptococcus pneumoniae (Strain JY2008, ATCC, Manassas, VA) was used in specific quantities over delineated time points.

Techniques: Cell Culture

Journal: Alcohol (Fayetteville, N.Y.)

Article Title: The Impact of Alcohol Use Disorders on Pulmonary Immune Cell Inflammatory Responses to Streptococcus pneumoniae

doi: 10.1016/j.alcohol.2018.08.016

Figure Lengend Snippet: Peripheral blood mononuclear cells (PBMCs) from non-AUD subjects (n=10) and AUD subjects (n=10) were cultured up to 42 hours, with and without the addition of 10 μg heat-killed S. pneumoniae (Sp). In some wells, N-acetylcysteine (NAC) was added after 18 hours in culture. Culture media was collected at the 18 hour and 42 hour time points for analysis of interferon (IFN)-γ, interleukin (IL)-1β, IL-6, and tumor necrosis factor (TNF)-α. (7A) With pneumococcal protein stimulation, IFNγ secretion by PBMCs rose significantly between the 18 and 42 hour time points (p=0.04). The addition of NAC at 18 hours during the 42-hour time course was associated with substantially less IFNγ secretion than that by non-NAC treated PBMCs (p=0.002 for both non-AUD and AUD subjects), with values that were significantly different than PBMCs at the 18 hour time point in non-AUD subjects only (p=0.02). (7B) With pneumococcal protein stimulation, IL-1β secretion by PBMCs did not rise significantly between the 18 and 42 hour time points (p=0.92). The addition of NAC at 18 hours during the 42-hour time course was associated with substantially less IL-1β secretion than that by non-NAC treated PBMCs at the 42 hour time point (p=0.002 for non-AUD subjects; p=0.02 for AUD subjects). Values among NAC-treated cells at the 42 hour time point approximated those in untreated cells at the 18 hour time point (p=ns). (7C) With pneumococcal protein stimulation, IL-6 secretion by PBMCs tended to rise between the 18 hour and 42 hour time points, but not significantly (p=0.09). The addition of NAC at 18 hours during the 42-hour time course was associated with substantially less IL-6 in cell culture supernatants than that found in supernatants from non-NAC treated PBMCs at the 42 hour time point (p=0.002 for both non-AUD and AUD subjects); values were not significantly different than PBMCs at the 18 hour time point for both subject types. (7D) With pneumococcal protein stimulation, TNFα secretion by PBMCs did not rise significantly between the 18 and 42 hour time points (p=0.44). The addition of NAC at 18 hours during the 42-hour time course was associated with substantially less TNFα in cell culture supernatants than in non-NAC treated PBMCs (p=0.002 for both non-AUD and AUD subjects); TNFα quantity in cell culture supernatant from pneumococcal protein -stimulated, NAC treated cells at 42 hours were significantly less than pneumococcal protein-stimulated PBMCs at the 18 hour time point among non-AUD subjects only (p=0.002). # indicates p≤0.02 between NAC and non-NAC treated conditions. & indicates p≤0.02 between 18-hour condition and NAC-treated, 42-hour condition.

Article Snippet: In some experiments, heat-killed Streptococcus pneumoniae (Strain JY2008, ATCC, Manassas, VA) was used in specific quantities over delineated time points.

Techniques: Cell Culture

Journal: Viruses

Article Title: Evaluation of a Lyophilized CRISPR-Cas12 Assay for a Sensitive, Specific, and Rapid Detection of SARS-CoV-2

doi: 10.3390/v13030420

Figure Lengend Snippet: Cross-reaction evaluation of Lyo-CRISPR SARS-CoV-2 kit with other respiratory pathogens.

Article Snippet: Streptococcus pneumoniae , ATCC ® 700669D-5 , 10 6 copies/mL , 0/3 , Negative , 3/3 , Valid.

Techniques: Concentration Assay, Virus

Journal: Investigative Ophthalmology & Visual Science

Article Title: Antibodies to Conserved Surface Polysaccharides Protect Mice Against Bacterial Conjunctivitis

doi: 10.1167/iovs.18-23795

Figure Lengend Snippet: Intraperitoneal administration of MAb to PNAG preinfection and postinfection reduces bacterial burdens and conjunctival pathology due to S. pneumoniae (D-39 and ATCC8) and S. aureus (LAC) after 48 hours of infection in A/J mice. Eighteen hours prior to infection and 4, 24, and 32 hours postinfection, 200 μg of the human IgG1 monoclonal antibody MAb F598 to PNAG or control IgG1 MAb F429 were injected IP. Mice (n = 8 per group) were euthanized at 48 hours postinfection and pathology scores (upper row) and CFU/conjunctiva (lower row) were determined. Symbols represent individual animals, black lines the median, and gray lines the 95% CI, and one-sided P values were determined by nonparametric t-tests.

Article Snippet: Streptococcus pneumoniae strains D-39 and ATCC8 were obtained from BEI Resources (Manassas, VA, USA) and grown in Todd-Hewitt Broth (BD Difco, Houston, TX, USA) with 1% added glucose statically in a 5% CO 2 incubator until they reached the mid-log phase of growth.

Techniques: Infection, Control, Injection

Journal: Investigative Ophthalmology & Visual Science

Article Title: Antibodies to Conserved Surface Polysaccharides Protect Mice Against Bacterial Conjunctivitis

doi: 10.1167/iovs.18-23795

Figure Lengend Snippet: Postinfection intraconjunctival administration of MAb to PNAG reduces bacterial burdens and conjunctival pathology due to S. pneumoniae (strains D-39 and ATCC8) and S. aureus (strain LAC) after 48 hours of conjunctival infection in A/J mice. A total of 10 μg of control IgG MAb (□) or MAb to PNAG (▵) were injected into the conjunctiva 4, 24, and 32 hours postinfection. Mice (n = 8 per group) were euthanized at 48 hours postinfection and pathology scores (upper row) and CFU/conjunctiva (lower row) were determined. Symbols represent individual animals, black lines the median, and gray lines the 95% CI, and one-sided P values were determined by nonparametric t-tests.

Article Snippet: Streptococcus pneumoniae strains D-39 and ATCC8 were obtained from BEI Resources (Manassas, VA, USA) and grown in Todd-Hewitt Broth (BD Difco, Houston, TX, USA) with 1% added glucose statically in a 5% CO 2 incubator until they reached the mid-log phase of growth.

Techniques: Infection, Control, Injection

Journal: Investigative Ophthalmology & Visual Science

Article Title: Antibodies to Conserved Surface Polysaccharides Protect Mice Against Bacterial Conjunctivitis

doi: 10.1167/iovs.18-23795

Figure Lengend Snippet: Postinfection topical administration of MAb to PNAG reduces bacterial burdens and conjunctival pathology due to S. pneumoniae (D-39 and ATCC8) and S. aureus (LAC) after 48 hours of infection in A/J mice. A total of 10 μg of control IgG MAb (□) or MAb to PNAG (▵) were administered topically onto conjunctiva 4, 24, and 32 hours postinfection. Mice (n = 7 to 8 per group) were euthanized at 48 hours post infection and pathology scores (upper row) and CFU/conjunctiva (lower row) determined. Symbols represent individual animals, black lines the median, and gray lines the 95% CI, and one-sided P values were determined by nonparametric t-tests.

Article Snippet: Streptococcus pneumoniae strains D-39 and ATCC8 were obtained from BEI Resources (Manassas, VA, USA) and grown in Todd-Hewitt Broth (BD Difco, Houston, TX, USA) with 1% added glucose statically in a 5% CO 2 incubator until they reached the mid-log phase of growth.

Techniques: Infection, Control

Journal: Investigative Ophthalmology & Visual Science

Article Title: Antibodies to Conserved Surface Polysaccharides Protect Mice Against Bacterial Conjunctivitis

doi: 10.1167/iovs.18-23795

Figure Lengend Snippet: Postinfection administration of MAb to PNAG reduces bacterial burdens and conjunctival pathology due to S. pneumoniae (D-39 and ATCC8) or S. aureus (LAC) in WT C57Bl/6, but not in RAG 1 KO or germ-free mice (GFM) after 48 hours. A total of 10 μg of control IgG MAb (□) or MAb to PNAG (▵) were injected into the conjunctiva 4, 24, and 32 hours postinfection. Mice (n = 5 per group) were euthanized at 48 hours postinfection and pathology scores (upper row) and CFU/conjunctiva (lower row) were determined. Symbols represent individual animals, bars the median, and error bars the 95% CI, and one-sided P values were determined by nonparametric t-tests. None of the differences between MAb to PNAG or control MAb in RAG 1 KO or GFM mice were significant (P > 0.22).

Article Snippet: Streptococcus pneumoniae strains D-39 and ATCC8 were obtained from BEI Resources (Manassas, VA, USA) and grown in Todd-Hewitt Broth (BD Difco, Houston, TX, USA) with 1% added glucose statically in a 5% CO 2 incubator until they reached the mid-log phase of growth.

Techniques: Control, Injection

Journal: Investigative Ophthalmology & Visual Science

Article Title: Antibodies to Conserved Surface Polysaccharides Protect Mice Against Bacterial Conjunctivitis

doi: 10.1167/iovs.18-23795

Figure Lengend Snippet: Postinfection administration of MAb to PNAG reduces conjunctival PMN infiltration due to S. pneumoniae (D-39 and ATCC8) and S. aureus (LAC) after 48 hours of conjunctival infection in A/J mice. A total of 10 μg of control IgG MAb or MAb to PNAG were injected into the conjunctiva 4, 24, and 32 hours postinfection in mice infected with S. pneumoniae D-39 or ATCC8 or S. aureus LAC. Control MAb-treated mice showed a large infiltrate of inflammatory cells and obvious edema in conjunctival areas, whereas mice treated with MAb to PNAG had only low levels of inflammatory cells and little edema. Arrows point to site of injection in the conjunctiva. Green bars: 10 μm. Boxed areas, when shown, indicated magnified area.

Article Snippet: Streptococcus pneumoniae strains D-39 and ATCC8 were obtained from BEI Resources (Manassas, VA, USA) and grown in Todd-Hewitt Broth (BD Difco, Houston, TX, USA) with 1% added glucose statically in a 5% CO 2 incubator until they reached the mid-log phase of growth.

Techniques: Infection, Control, Injection

Journal: Investigative Ophthalmology & Visual Science

Article Title: Antibodies to Conserved Surface Polysaccharides Protect Mice Against Bacterial Conjunctivitis

doi: 10.1167/iovs.18-23795

Figure Lengend Snippet: Postinfection administration of MAb to PNAG reduces conjunctival myeloperoxidase due to S. pneumoniae (D-39 and ATCC8) and S. aureus (LAC) after 48 hours of conjunctival infection in A/J mice. A total of 10 μg of control IgG MAb (□) or MAb to PNAG (▵) were injected into the conjunctiva 4, 24, and 32 hours postinfection with S. pneumoniae (D-39, ATCC8) or S. aureus LAC. Mice were euthanized at 48 hours postinfection and MPO/conjunctiva was determined. Symbols represent individual animals, black lines the medians, and gray lines the 95% CI, and P values were determined by one-sided nonparametric t-tests.

Article Snippet: Streptococcus pneumoniae strains D-39 and ATCC8 were obtained from BEI Resources (Manassas, VA, USA) and grown in Todd-Hewitt Broth (BD Difco, Houston, TX, USA) with 1% added glucose statically in a 5% CO 2 incubator until they reached the mid-log phase of growth.

Techniques: Infection, Control, Injection