i scapularis tick cell line ise6  (ATCC)

Journal: iScience

Article Title: GW4869 inhibitor affects vector competence and tick-borne flavivirus acquisition and transmission by blocking exosome secretion

doi: 10.1016/j.isci.2024.110391

Figure Lengend Snippet: Efficiency of LGTV dissemination within I. scapularis nymphal ticks is affected upon GW4869 treatment (A–C) LGTV loads in midgut (MG), salivary glands (SGs) or carcass (C) of nymphs that were simultaneously exposed to LGTV (1 MOI) and either mock (1.5% DMSO) or GW4869 (150 μM) and incubated for 0 (A), 3 (B) or 17 (C) days post infection (DPI), respectively. (D and E) LGTV loads in MG, C and SGs of nymphs that were pretreated with either mock/GW4869 followed by LGTV infection and incubated for 3 (C) or 17 (D) days p.i., respectively. LGTV loads were normalized to total RNA. Circles denote the mock-treated group, whereas squares represent the GW4869-treated group. Each circle/square represents a pool of either two (A–C) or three (D and E) ticks in the simultaneous or pretreatment groups, respectively. p value less than 0.05 is considered statistically significant and ns indicates not significant.

Article Snippet: The I. scapularis ISE6 tick cell line was purchased from ATCC and grown in L15B300 medium prepared from Leibovitz’s L-15 Medium powder with 5% tryptose phosphate broth, 5% heat-inactivated FBS, and 0.1% bovine lipoprotein concentrate, pH 7.2.

Techniques: Incubation, Infection

Journal: iScience

Article Title: GW4869 inhibitor affects vector competence and tick-borne flavivirus acquisition and transmission by blocking exosome secretion

doi: 10.1016/j.isci.2024.110391

Figure Lengend Snippet: LGTV loads and EVs released from salivary glands isolated from I. scapularis female ticks are reduced upon GW4869 treatment (A) QRT-PCR showing LGTV loads in midgut (MG), carcass (C), and salivary glands (SGs) of adult ticks that were simultaneously exposed to LGTV (1 MOI) and to either mock (1.5% DMSO) or GW4869 (150 μM) and incubated for 3 days post-infection. (B) LGTV loads shown from SGs-derived EVs isolated from female ticks incubated at 3 days post LGTV infection with simultaneous treatment with mock/GW4869. (C) Concentration of EVs-derived from SGs isolated from female ticks incubated for 3 days post LGTV infection and mock/GW4869 treatment is shown. (D and E) Graphical representation showing the quantification of EVs-derived from SGs isolated from female ticks after 3 days post LGTV infection and mock/GW4869 treatment. Four-five independent replicates were considered for QRT-PCR or for determining EVs concentration. LGTV loads were normalized to total RNA. Circles denote mock-treated group, whereas squares represent the GW4869-treated group. Each circle/square represents tissues from one adult tick (in A and B) and salivary glands collected from two female ticks (C), respectively. p value less than 0.05 is considered statistically significant and ns indicates not significant.

Article Snippet: The I. scapularis ISE6 tick cell line was purchased from ATCC and grown in L15B300 medium prepared from Leibovitz’s L-15 Medium powder with 5% tryptose phosphate broth, 5% heat-inactivated FBS, and 0.1% bovine lipoprotein concentrate, pH 7.2.

Techniques: Isolation, Quantitative RT-PCR, Incubation, Infection, Derivative Assay, Concentration Assay

Journal: iScience

Article Title: GW4869 inhibitor affects vector competence and tick-borne flavivirus acquisition and transmission by blocking exosome secretion

doi: 10.1016/j.isci.2024.110391

Figure Lengend Snippet: GW4869 treatment reduces molting efficiency and survival rates of LGTV-infected ticks (A) Molting rates of untreated uninfected or LGTV-infected or GW4869 (50 μM)-treated and LGTV-infected I. scapularis larvae into nymphs is shown. (B) Total survival rates of untreated uninfected or LGTV-infected or GW4869 (50 μM)-treated and LGTV-infected larvae molting into nymphs are shown. (C) Total survival rates of mock (1.5% DMSO) or GW4869 (150 μM)- treated and LGTV-infected nymphs molting into adults are shown. (D) LGTV loads in adult ticks molted from mock/GW4869-treated and LGTV-infected nymphs is shown. LGTV loads were normalized to total RNA. Circles denote mock-treated group, whereas squares represent the GW4869-treated group. Each circle/square represents one tick. p value less than 0.05 is considered statistically significant and ns indicates not significant. (E) Model showing the effects of the GW4869 treatment of ticks on LGTV transmission to the vertebrate host. Graphical representation shows that LGTV-infected and GW4869-treated (150 μM) ticks are hampered in blood feeding (with reduced tick body weights). GW4869-treated and LGTV-infected ticks fed partially showing reduced feeding efficiency and decreased viral transmission into the vertebrate host. Treatment of GW4869 affected EVs release that corelated with significant reduction in LGTV loads from these ticks. The GW4869 treatment resulted in inefficient tick feeding, lower EVs biogenesis, decreased viral loads during acquisition and transmission, and reduced molting/survival rates. The combination of all these factors eventually affected vector fitness and competence.

Article Snippet: The I. scapularis ISE6 tick cell line was purchased from ATCC and grown in L15B300 medium prepared from Leibovitz’s L-15 Medium powder with 5% tryptose phosphate broth, 5% heat-inactivated FBS, and 0.1% bovine lipoprotein concentrate, pH 7.2.

Techniques: Infection, Transmission Assay, Plasmid Preparation

Journal: iScience

Article Title: GW4869 inhibitor affects vector competence and tick-borne flavivirus acquisition and transmission by blocking exosome secretion

doi: 10.1016/j.isci.2024.110391

Figure Lengend Snippet:

Article Snippet: The I. scapularis ISE6 tick cell line was purchased from ATCC and grown in L15B300 medium prepared from Leibovitz’s L-15 Medium powder with 5% tryptose phosphate broth, 5% heat-inactivated FBS, and 0.1% bovine lipoprotein concentrate, pH 7.2.

Techniques: Virus, Saline, SYBR Green Assay, Transfection, cDNA Synthesis, Gel Extraction, Software, Imaging