Combined deficiency of coagulation factors V and VIII (F5F8D) is an autosomal recessive bleeding disorder caused by loss-of-function mutations in either LMAN1 or MCFD2. The latter genes encode 2 ...components of a mammalian cargo receptor that facilitates secretion of coagulation factor V (FV) and factor VIII (FVIII) from the endoplasmic reticulum (ER) to the Golgi via coat protein complex II vesicles. F5F8D patients exhibit FV and FVIII levels that are ∼10% to 15% of normal. We report herein a comparative analysis for a series of murine Lman1 alleles. Consistent with previous reports, mice completely deficient in LMAN1 (Lman1−/−) exhibit ∼50% FV and FVIII levels. In contrast, mice carrying a hypomorphic Lman1 allele (Lman1cgt/cgt) that expresses ∼6% to 8% of wild-type Lman1 mRNA levels exhibit intermediate plasma FV and FVIII reductions (∼70% of wild-type levels). Lman1−/− mice exhibit ER accumulation of another LMAN1 cargo, alpha-1 antitrypsin (A1AT), with an intermediate level of A1AT ER retention observed in Lman1cgt/cgt mice. Finally, the previously reported strain-specific, partially penetrant, perinatal lethality of LMAN1-deficient mice (Lman1gt1/gt1) was confirmed in Lman1−/− mice, although it was not observed in Lman1cgt/cgt mice. Taken together, these results show a dose-dependent effect of residual LMAN1 on the secretion of its cargo proteins. The results also suggest that human subjects with hypomorphic LMAN1 mutations might present with mild bleeding phenotypes resulting from more modest reductions in FV and FVIII, which could be missed by routine clinical evaluation. Finally, these findings suggest that therapeutic targeting of LMAN1 to reduce FV and FVIII as an anticoagulant strategy may only require partial inhibition of LMAN1 function.
•There is a dose–response relationship between Lman1 expression level and secretion levels of LMAN1 cargoes in mice (FV, FVIII, and A1AT).•Therapeutic targeting of LMAN1 to reduce FV and FVIII as an anticoagulant strategy may only require partial inhibition of LMAN1 function.
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Cross-presentation initiates immune responses against tumors and viral infections by presenting extracellular antigen on MHC I to activate CD8+ T cell-mediated cytotoxicity. In vitro studies in ...dendritic cells (DCs) established SNARE protein SEC22B as a specific regulator of cross-presentation. However, the in vivo contribution of SEC22B to cross-presentation has not been tested. To address this, we generated DC-specific Sec22b knockout (CD11c-Cre Sec22bfl/fl) mice. Contrary to the paradigm, SEC22B-deficient DCs efficiently cross-present both in vivo and in vitro. Although in vitro small hairpin RNA (shRNA)-mediated Sec22b silencing in bone-marrow-derived dendritic cells (BMDCs) reduced cross-presentation, treatment of SEC22B-deficient BMDCs with the same shRNA produced a similar defect, suggesting the Sec22b shRNA modulates cross-presentation through off-target effects. RNA sequencing of Sec22b shRNA-treated SEC22B-deficient BMDCs demonstrated several changes in the transcriptome. Our data demonstrate that contrary to the accepted model, SEC22B is not necessary for cross-presentation, cautioning against extrapolating phenotypes from knockdown studies alone.
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•SEC22B is not necessary for cross-presentation of ovalbumin in vivo or in vitro•Off-target effects from Sec22b-targeting shRNA reduce cross-presentation•RNA sequencing identifies putative cross-presentation mediators
SEC22B has been characterized as a crucial mediator of antigen cross-presentation. Using DC-specific Sec22b knockout mice, Wu et al. demonstrate that SEC22B is not necessary for cross-presentation and that the Sec22b shRNA sequences used in prior studies impact cross-presentation through off-target effects. They identify potential cross-presentation mediators for future investigation.
The highly conserved SNARE protein SEC22B mediates diverse and critical functions, including phagocytosis, cell growth, autophagy, and protein secretion. However, these characterizations have thus ...far been limited to in vitro work. Here, we expand our understanding of the role Sec22b plays in vivo. We utilized Cre-Lox mice to delete Sec22b in three tissue compartments. With a germline deletion of Sec22b, we observed embryonic death at E8.5. Hematopoietic/endothelial cell deletion of Sec22b also resulted in in utero death. Notably, mice with Sec22b deletion in CD11c-expressing cells of the hematopoietic system survive to adulthood. These data demonstrate Sec22b contributes to early embryogenesis through activity both in hematopoietic/endothelial tissues as well as in other tissues yet to be defined.
Acquired thrombotic thrombocytopenic purpura is a rare blood disorder with a high early mortality rate, if untreated. Standard of care plasma exchange and glucocorticoids have dramatically improved ...survival. However, additional advancements are necessary to further decrease mortality. Caplacizumab-yhdp (Cablivi®) is the first Food and Drug Administration-approved treatment indicated for adult patients with acquired thrombotic thrombocytopenic purpura, in combination with plasma exchange and immunosuppressive therapy. However, there are considerable risks associated with the use of caplacizumab and they must be weighed against the benefits of the medication.
The changing face of neurosyphilis Chahine, Lama M.; Khoriaty, Rami N.; Tomford, Walton J. ...
International Journal of Stroke,
04/2011, Letnik:
6, Številka:
2
Journal Article, Book Review
Recenzirano
The incidence of syphilis has increased over the past decade, particularly among HIV-positive patients, and the presenting clinical features have changed since the beginning of the HIV epidemic. The ...clinical manifestations of neurosyphilis are protean, and include acute stroke. In patients with HIV, the diagnosis and treatment of neurosyphilis is challenging. We review the clinical presentation, pathophysiology, and treatment of neurosyphilis, with emphasis on neurosyphilis in the HIV population, and neurosyphilis as a cause of acute stroke.
During the analysis of a whole genome ENU mutagenesis screen for thrombosis modifiers, a spontaneous 8 base pair (bp) deletion causing a frameshift in exon 27 of the Nbeal2 gene was identified. ...Though initially considered as a plausible thrombosis modifier, this Nbeal2 mutation failed to suppress the synthetic lethal thrombosis on which the original ENU screen was based. Mutations in NBEAL2 cause Gray Platelet Syndrome (GPS), an autosomal recessive bleeding disorder characterized by macrothrombocytopenia and gray-appearing platelets due to lack of platelet alpha granules. Mice homozygous for the Nbeal2 8 bp deletion (Nbeal2gps/gps) exhibit a phenotype similar to human GPS, with significantly reduced platelet counts compared to littermate controls (p = 1.63 x 10-7). Nbeal2gps/gps mice also have markedly reduced numbers of platelet alpha granules and an increased level of emperipolesis, consistent with previously characterized mice carrying targeted Nbeal2 null alleles. These findings confirm previous reports, provide an additional mouse model for GPS, and highlight the potentially confounding effect of background spontaneous mutation events in well-characterized mouse strains.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Telomerase catalyzes chromosome end replication in stem cells and other long-lived cells. Mutations in telomerase or telomere-related genes result in diseases known as telomeropathies. Telomerase is ...recruited to chromosome ends by the ACD/TPP1 protein (TPP1 hereafter), a component of the shelterin complex that protects chromosome ends from unwanted end joining. TPP1 facilitates end protection by binding shelterin proteins POT1 and TIN2. TPP1 variants have been associated with telomeropathies but remain poorly characterized in vivo. Disease variants and mutagenesis scans provide efficient avenues to interrogate the distinct physiological roles of TPP1. Here, we conduct mutagenesis in the TIN2- and POT1-binding domains of TPP1 to discover mutations that dissect TPP1's functions. Our results extend current structural data to reveal that the TPP1-TIN2 interface is more extensive than previously thought and highlight the robustness of the POT1-TPP1 interface. Introduction of separation-of-function mutants alongside known TPP1 telomeropathy mutations in mouse hematopoietic stem cells (mHSCs) lacking endogenous TPP1 demonstrated a clear phenotypic demarcation. TIN2- and POT1-binding mutants were unable to rescue mHSC failure resulting from end deprotection. In contrast, TPP1 telomeropathy mutations sustained mHSC viability, consistent with their selectively impacting end replication. These results highlight the power of scanning mutagenesis in revealing structural interfaces and dissecting multifunctional genes.
Oncology boards flash review Worden, Francis P; Khoriaty, Rami N
2013., 2013, 20130101, 20130506, 2013-05-06
eBook
Oncology Boards Flash Review is a question-and-answer book designed to summarize the most important facts one needs to know for the medical oncology boards, including the most up-to-date information ...on well-established chemotherapy regimens for a variety of malignancies. Coverage extends across the range of major hematologic malignancies and solid tumors, and includes reviews of pharmacology, biostatistics, genetics and tumor biology, and survivorship and palliative care. The book covers all of the topics listed by the American Board of Internal Medicine as essential material for the Medical Oncology Board Examination. Each chapter is written by a fellow and edited by expert faculty and clinicians, and Flash Review includes key points summarized in bullet point form for easy recall. Question- based headings enable the reader to systematically review and assess his knowledge. This study guide distills the must-know points of medical oncology into a single source for targeted board review, self-assessment, or handy quick reference. Oncology Boards Flash Review features: * Must-know points of medical oncology * Coverage of hematologic malignancies, solid tumors, pharmacology, biostatistics, genetics and tumor biology, and survivorship and palliative care * Key points summarized in bullet form for easy recall * Each chapter written by a fellow and edited by expert faculty and clinicians * Useful tool for board review, self-assessment or, handy quick reference
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