Tamoxifen-induced recombinase activity of Cre-ER^T2 in the lung tissues of SPC-Cre-ER^T2/TSC1^fx/fx transgenic mice.

<p>DNAs from the lung tissues of SPC-Cre-ER^T2/TSC1^fx/fx transgenic mice treated with vehicle or tamoxifen were examined by PCR to detect <i>TSC1</i> deletion as an indication of Cre-ER^T2 recombinase activity. M, DNA marker; +, a transgenic mouse with <i>TSC1</i> deletion; -, C57BL/6J mouse. Other lanes, offspring from F42 and F67 founders treated with vehicle (vehi) or tamoxifen (tam).</p

Image1_Case Report: ALK rearranged locally advanced lung adenocarcinoma showing inconsistent radiographic findings and pathological responses during neoadjuvant alectinib therapy.JPEG

Alectinib has been approved as first-line treatment for anaplastic lymphoma kinase (ALK)-positive non-small cell lung carcinoma. Oncologists are also exploring the possibility of applying alectinib in the perioperative period. Here, we present a patient with locally advanced lung adenocarcinoma associated with EML4-ALK fusion mutation, who received neoadjuvant chemotherapy and alectinib treatment, and then underwent thoracoscopic left lower lung lobectomy. The patient initially received eight chemotherapy cycles and achieved partial remission. After eight cycles of chemotherapy, the lymph nodes in the hilar region again enlarged. The patient was then switched to 4 months of alectinib therapy, but no significant lesion changes were detected on imaging during this period. This raised the question of whether the patient developed alectinib resistance. The pathological findings of the postoperative lung lobe specimens indicated extensive necrosis in the tumor area with no residual tumor cells and massive chronic inflammatory cell infiltration around the tumor area, confirming inconsistency between the imaging findings and pathological results. Multi-point tumor specimen sampling was postoperatively performed. Tumor immune-related gene expression was detected in the sample with the help of the PanCancer IO360™ panel based on the nCounter platform. This is a rare case of a patient who was treated with neoadjuvant alectinib and had paradoxical radiographic findings and pathological responses. The possibility that intratumoral immune heterogeneity was responsible for this phenomenon has been discussed. Based on the findings, it is argued that the pathological response should be an important basis for assessing the effectiveness of neoadjuvant alectinib therapy.</p

Generation of SPC-Cre-ER^T2 mice.

<p>A) Schematic map of SPC-Cre-ER^T2 expression cassette. HSPC-P, human surfactant protein C promoter; Cre-ER^T2, Cre coding sequence fused with a tamoxifen-inducible estrogen receptor. pA, a polyA sequence from SV40 virus. Cre-F primer binding sites, 561–579 bp of Cre-ER^T2 transgene; Cre-R primer binding sites, 976–997 bp of Cre-ER^T2 transgene. The map is drawn in scale. B) Screening SPC-Cre-ER^T2 transgenic mice using PCR. Genomic DNA from each mouse tail was used as template to specifically PCR-amplify the Cre-ER^T2 transgene. M, DNA marker; +, SPC-Cre-ER^T2 plasmid DNA control; -, water control; F8, F13, F16, F42, F67 are five representative founder transgenic mice generated by microinjection of SPC-Cre-ER^T2 expression cassette into fertilized embryos. C) Cre-ER^T2 fusion proteins were detected using Western blot in lung tissues of SPC-Cre-ER^T2 transgenic mice. Lane 1, C57BL/6J mouse; Lane 2, an offspring of F42 founder without Cre-ER^T2 transgene when genotyped using PCR; Lane 3, 4, 5, offspring of F42 founder; Lane 6, 7, 8, offspring of F67 founder. Notice the variable expression levels of Cre-ER^T2 in offspring from the same founder.</p

Tamoxifen-inducible and tissue-specific recombinase activity of Cre-ER^T2 in SPC-Cre-ER^T2/ROSA26R transgenic mice.

<p>A) β-galactosidase activity was detected in the lung alveolar epithelia from a SPC-Cre-ER^T2/ROSA26R transgenic mouse receiving tamoxifen treatment. a & b, lung tissue sections from ROSA26R mice (without Cre-ER^T2 transgene) receiving vehicle (a) or tamoxifen (b); c & d, lung tissue sections from SPC-Cre-ER^T2/ROSA26R transgenic mice receiving vehicle (c) or tamoxifen (d). B) Endogenous β-galactosidase activity was found in lung bronchial epithelia of ROSA26R mouse receiving vehicle (a) and SPC-Cre-ER^T2/ROSA26R transgenic mouse after tamoxifen treatment (b). C) X-gal stained lung tissues of SPC-Cre-ER^T2/ROSA26R transgenic mouse receiving tamoxifen were also immune-stained for proSP-C, an alveolar type II cell-specific marker. Arrows indicate three representative alveolar type II cells co-expressing proSP-C (brown) and β-galactosidase (blue). D) β-galactosidase activity was detected only in the lung alveolar epithelium, but not in other organs from a SPC-Cre-ER^T2/ROSA26R transgenic mouse receiving tamoxifen treatment. a, heart; b, liver; c, kidney; d, intestine; e, lung. All scale bars in this figure equal 100 µm.</p

Omeprazole Alleviates <i>Aristolochia manshuriensis Kom</i>-Induced Acute Nephrotoxicity

<div><p><i>Aristolochia manshuriensis Kom</i> (AMK) is a member of the <i>Aristolochiaceae</i> family and is a well-known cause of aristolochic acid (AA) nephropathy. In this study, we investigated the potential of omeprazole (OM) to alleviate AMK-induced nephrotoxicity. We found that OM reduced mouse mortality caused by AMK and attenuated AMK-induced acute nephrotoxicity in rats. OM enhanced hepatic Cyp 1a1/2 and renal Cyp 1a1 expression in rats, as well as CYP 1A1 expression in human renal tubular epithelial cells (HKCs). HKCs with ectopic CYP 1A1 expression were more tolerant to AA than the control cells. Therefore, OM may alleviate AMK-mediated acute nephrotoxicity through induction of CYP 1A1. We suggest that the coadministration of OM might be beneficial for reducing of AA-induced nephrotoxicity.</p></div

Additional file1: of miR-182 suppresses invadopodia formation and metastasis in non-small cell lung cancer by targeting cortactin gene

Table S1. Basic information of 55 patients with NSCLC. (PDF 64 kb

Decreased Akt-FoxO1/FoxO3a phosphorylation of Tsc1 KO naïve CD8⁺T cells in response to IL-7.

<p>The expression of CD122, CD127 and CD95 on naïve CD8⁺T cells was determined by gating CD8⁺CD44^low cells. Comparable surface CD122, CD127 (<b>A</b>) and CD95 (<b>B</b>) expression in Tsc1 KO naïve CD8⁺ T cells. The dash-dot line represents staining with an isotype control antibody. The histograms (gray) represent WT whereas the open histograms with solid line show Tsc1 KO staining pattern. Cells are gated with CD8⁺CD44^low population. Representative data are shown from one of two separate experiments, with three mice in each group. (<b>C</b>) Western blot analysis of mTORC2-Akt-FoxO1/FoxO3a-Bim axis in naïve CD8⁺ T cells after stimulation with IL-7. Sorted naïve CD8⁺T cells were cultured with IL-7 in the presence of Rapa or not for 24 hrs. The freshly isolated WT or Tsc1 KO naïve CD8⁺T cells were used as a control. One representative is shown from two or three separate experiments.</p

TSC1 regulates naïve CD8⁺ T cell survival to IL-7 or IL-15 in a rapamycin-insensitive manner.

<p>Sorted WT and Tsc1 KO mouse naïve CD8⁺CD44^low T cells were cultured in medium alone or supplemented with IL-15 or IL-7(in the presence or absence of Rapa) for 24 hrs or the below indicated time points. (<b>A</b>) Significantly decreased live cell number of Tsc1 KO naïve CD8⁺T cells than WT naïve CD8⁺T cells in the presence of IL-15 or IL-7 for 24 hours. Live cells were determined by trypan blue exclusion assay. (<b>B</b>) Tsc1 KO naïve CD8⁺ T cells showed severe atrophy in the presence of IL-7 for 24 hrs. (<b>C</b>) One representative of PI staining in gated CD8⁺T cells after culture with or without IL-7 and IL-15. (<b>D</b>) Percentage of cell death was measured by PI staining and statistically analyzed. The above data were one representative of three separate experiments, with three wells in each group. (<b>E</b>) Representative staining of CD45.2 and CD8 for pLN cells of CD45.1⁺ host in the presence or absence of Rapa 7 days after transfer. Either sorted WT or Tsc1 KO CD45.2⁺ naïve CD8⁺ T cells were adoptively transferred into sublethally irradiated(4 Gy) CD45.1⁺ host and the donor cells were determined by CD8 and CD45.2 staining. (<b>F</b>) Significantly decreased percentage and cell number of Tsc1 KO CD45.2⁺ naïve CD8⁺ T cells in spleen of sublethally irradiated (4 Gy) CD45.1⁺ host in the presence or absence of Rapa. (<b>G</b>) Significantly decreased percentage and cell number of Tsc1 KO CD45.2⁺ naïve CD8⁺ T cells in pLNs of sublethally irradiated (4 Gy) CD45.1⁺ host in the presence or absence of Rapa. Data were shown as Mean±SD (3 mice each group). *p<0.05; **p<0.01; ***p<0.001 compared with the indicated groups.</p

Omeprazole protects rats against <i>Aristolochia manshuriensis Kom</i>-mediated renal injury.

<p>Alterations in (A) female and (B) male body weight, (C) female and (D) male Kidney weight body weight ratio (KW/BW), (E) female and (F) male serum BUN, and (G) female and (H) male serum CRE in the four rat cohorts. qPCR analysis of (I) female and (J) male renal <i>ngal</i> mRNA levels in the four rat cohorts. Urine Ngal levels of (K) female and (L) males in each cohort. CTL, control; AMK, <i>Aristolochia manshuriensis Kom</i>; OM, omeprazole; O+A, omeprazole and AMK. Data are expressed as mean ± SEM. *P<0.05 <i>vs</i>. CTL cohort; **P<0.01 <i>vs</i>. CTL cohort; ***P<0.001 <i>vs</i>. CTL cohort; ^# P<0.05 <i>vs</i>. AMK cohort; ^## P<0.01 <i>vs</i>. AMK cohort; ^### P<0.001 <i>vs</i>. AMK cohort. n = 6~7.</p

Naïve CD8⁺ but not CD4⁺ T cells were significantly decreased in the periphery of Tsc1 KO mice.

<p>The peripheral T cell subsets of WT and Tsc1 KO mice were detected using FCM 4–5 wks after birth. <b>A.</b> Total thymic cell number and thymocyte subset profile of Tsc1 KO mice were identical as WT littermates. <b>B.</b> Total cell number of spleen, pLNs and mLNs of WT and Tsc1 KO mice. <b>C.</b> The percentage of CD8⁺T but not CD4⁺T cells decreased dramatically in all peripheral lymphoid organs of Tsc1 KO mice. <b>D.</b> The FACS profile analysis of CD62L, CD45RB and CD44 on pLN CD4⁺ T cells was evaluated. The frequency of naïve CD62L^hi, CD45RB^hi and CD44^low CD4⁺T cells in spleen, pLN and mLN of Tsc1 KO mice was summarized. <b>E.</b> The FACS profile analysis of CD62L, CD45RB and CD44 on the gated pLN CD8⁺ T cells was evaluated. The frequency of naïve CD62L^hi, CD45RB^hi and CD44^low CD8⁺T cells in spleen, pLN and mLN of Tsc1 KO mice was summarized. *p<0.05; **p<0.01; ***p<0.001. Data were shown as Mean±SD (N = 6).</p