Generation of two hiPSCs lines of two patients carrying truncating mutations in the dimerization domain of filamin C.

Here we introduce the human induced pluripotent stem cell lines (hiPSCs), HIMRi004-A and HIMRi005-A from dermal fibroblasts of a 48-year-old female (HIMRi004-A) carrying missense mutation that translate to the first described filamin C isoform p.W2710X and from a 56-year-old female (HIMRi005-A) carrying a recently described mutation in the same domain p.Y2704X. Both lines are generated via lentiviral expression of OCT4, SOX2, KLF4 and c-MYC. The lines display a typical embryonic stem cell-like morphology, express pluripotency markers, retain a normal karyotype (46, XX) and have the differentiation capacity in all three germ layers. The two lines can be used to elucidate the pathomechanisms of FLNC myofibrillar myopathies and to develop novel therapeutic options.

Generation of two human iPSC lines (HIMRi002-A and HIMRi003-A) derived from Caveolinopathy patients with rippling muscle disease.

Here we introduce the human induced pluripotent stem cell lines (hiPSCs), HIMRi002-A and HIMRi003-A, generated from cultured dermal fibroblasts of 61-year-old (HIMRi002-A) and 38-year-old (HIMRi003-A) female patients, carrying a known heterozygous pathogenic variant (p.A46T) in the Caveolin 3 (CAV3) gene, via lentiviral expression of OCT4, SOX2, KLF4 and c-MYC. HIMRi002-A and HIMRi003-A display typical embryonic stem cell-like morphology, carry the p.A46T CAV3 gene mutation, express several pluripotent stem cell markers, retain normal karyotype (46, XX) and can differentiate in all three germ layers. We postulate that the HIMRi002-A and HIMRi003-A iPSC lines can be used for the characterization of CAV3-associated pathomechanisms and for developing new therapeutic options.

Generation of a human iPSC line (HIMRi001-A) from a patient with filaminopathy.

Here we introduce the human induced pluripotent stem cell (hiPSC) line HIMRi001-A generated from cultured dermal fibroblasts of a 60-year-old male patient with a myofibrillar myopathy, carrying a heterozygous c.4984C > T [p.Q1662X] mutation in the filamin C (FLNC)-gene, via lentiviral expression of OCT4, SOX2, KLF4 and c-MYC. HIMRi001-A displays typical embryonic stem cell-like morphology, carries the c.4984C > T FLNC gene mutation, expressed several pluripotent stem cell makers, retained normal karyotype (46, XY) and holds the potential to differentiate in all three germ layers. We postulate that HIMRi001-A can be used for the elucidation of FLNC-associated pathomechanisms and for developing new therapeutic options.

[Detection of HFRSV in Eulaelaps shanghaiensis and Ornithonyssus bacoti by using in situ hybridization].

AIM: To provide molecular biological evidence of transmission of hemorrhogic fever with renal syndrome virus (HFRSV) by gamasid mites, Eulaelaps shanghaiensis and Ornithonyssus bacoti. METHODS: Frozen sections of the gamasid mites 10 days and more than 100 days after biting suckling mice inoculated with HFRSV were in situ hybridized with dig-labelled HFRSV cDNA probes. RESULTS: RNA was detected in frozen sections of Eulaelaps shanghaiensis, after biting suckling mice inoculated with Hantaan (76-118) and Seoul (UR) virus, respectively. Most of the fine granules of the virus RNA were located in the nuclei, cytoplasm and nuclear membrane of cells of brain cortex, caeca and genitalia of the mites. In situ hybridization results showed that 17 of 31 mites in Hantaan group and 10 of 23 mites in Seoul group were positive. The virus RNA was still detected in tissues of the mites on d132 for Hantaan group and on d102 for Seoul group after infection, respectively. Among 20 Ornithonyssus bacoti detected 12 were positive on d10 after biting suckling mice inoculated with Hantaan virus, and the virus RNA was mainly found in the cells of genitalia (Figs. 1-13). CONCLUSION: Both Eulaelaps shanghaiensis and Ornithonyssus bacoti could be infected with HFRSV by biting HFRSV-positive mice. E. shanghaiensis could be infected with both Hantaan and Seoul virus, and the two types of HFRSV were found to be maintained in the mites for 132 days and 102 days, respectively. These confirmed that E. shanghaiensis and O. bacoti are suitable vectors and reservoirs of both Hantaan and Seoul virus and might play an important role in the cross transmission of the two types of HFRSV.

[Studies on the experimental transmission of Rattus-borne Hantavirus by Ornithonyssus bacoti].

AIM: To demonstrate the role of rat mite (Ornithonyssus bacoti) in the transmission of Rattus-borne hemorrhagic fever with renal syndrome (HFRS). METHODS: In the transmission experiments, about 100 O. bacoti per pool were isolated and placed in a jar, unfed for 4 d at 23 +/- 1 degrees C. Suckling Wistar rats inoculated with Hantavirus strain Z45 or Seoul virus strain UR were placed in each jar for free attack by the mites for 12 hours. After 14 d the normal suckling Wistar rats were bitten by the mites. Fifteen days later, the lung tissues and sera of the infected rats were collected and detected for Hantaviral antigen by indirect fluorescent antibody technique (IFAT). For demonstration of the infection of O. bacti with Rattus-borne Hantavirus PCR technique was applied to detect Rattus-borne Hantaviral RNA. RESULTS: Sukling Wistar rats inoculted Hantavirus strain Z45 or Seoul virus strain UR were bitten by O. bacoti and then these mites were fed on 4 and 5 of normal suckling rats in each jar, respectively. The antigens of Hantavirus strain Z45 were positive in all the lungs of the normal rats bitten by the mites, the sera titers of the rats were from 1:10 to 1:40. The antigens were positive in 3 of the 4 rats, the sera titers were from 1:20 to 1:40. Both of the viruses could be maintained in O. bacoti for 22 days. The blocking test showed when 1:30 Hantavirus immunosera were exposed to the lung samples and then reacted with the sera from the patients with HFRS, all the specific fluorescence reactions of the samples were blocked, whereas the control group including the normal rat lung tissues and sera were all negative (Fig. 1). CONCLUSION: O. bacoti might play a role as the vector of HFRS and a reservoir host as well.