Stem cell-mediated transfer of a human artificial chromosome ameliorates muscular dystrophy

In contrast to conventional gene therapy vectors, human artificial chromosomes (HACs) are episomal vectors that can carry large regions of the genome containing regulatory elements. So far, HACs have not been used as vectors in gene therapy for treating genetic disorders. Here, we report the amelioration of the dystrophic phenotype in the mdx mouse model of Duchenne muscular dystrophy (DMD) using a combination of HAC-mediated gene replacement and transplantation with blood vessel-associated stem cells (mesoangioblasts). We first genetically corrected mesoangioblasts from dystrophic mdx mice with a HAC vector containing the entire (2.4 Mb) human dystrophin genetic locus. Genetically corrected mesoangioblasts engrafted robustly and gave rise to many dystrophin-positive muscle fibers and muscle satellite cells in dystrophic mice, leading to morphological and functional amelioration of the phenotype that lasted for up to 8 months after transplantation. Thus, HAC-mediated gene transfer shows efficacy in a preclinical model of DMD and offers potential for future clinical translation.     

Multiple IgG4-related sclerosing lesions in the maxillary sinus, parotid gland and nasal septum

IgG4-related sclerosing disease is recognized as a distinct clinicopathological entity. It is well known that this disease can occur in the salivary, lacrimal and pituitary glands, in the head and neck region. The nasal cavity is an extremely rare site of involvement of IgG4-related sclerosing disease. Herein is reported a case of multiple IgG4-related sclerosing lesions in the maxillary sinus, parotid gland and nasal septum. A 73-year-old Japanese man presented with nasal obstruction and tumors of the right maxillary sinus and parotid gland were detected, after which resections of these tumors were performed. One year after the last surgery, he noted swelling of the nasal septum, and the tumor was resected. These three tumors had similar histopathology, such as conspicuous fibrosclerotic changes with dense lymphoplasmacytic infiltration and occasional obliterative phlebitis. Immunohistochemistry indicated abundant IgG4-positive plasma cell infiltration and high ratios of IgG4-positive/IgG-positive plasma cells (>70%) in all three lesions. The diagnosis of multiple IgG4-related sclerosing lesions was made. The present case suggests that IgG4-related sclerosing lesion can occur in the maxillary sinus and nasal septum, and represents an extension of the spectrum of IgG4-related sclerosing disease.     

Preferential expression of RB1-inducible coiled-coil 1 in terminal differentiated musculoskeletal cells

RB1-inducible coiled-coil 1 (RB1CC1) is a nuclear DNA-binding protein that can induce RB1 (retinoblastoma 1) expression. RB1CC1 is abundantly expressed in human musculoskeletal and cultured osteosarcoma cells. The present study analyzed the expression of RB1CC1 and RB1 in osteosarcoma cells and in musculoskeletal cells of human embryos to evaluate the contribution of both genes to the maturational process of musculoskeletal cells. The amount of RB1CC1 message was closely related to RB1 expression in various osteosarcoma cell lines. RB1CC1 expression was difficult to detect in immature proliferating chondroblasts or myogenic cells in human embryos, but became obvious and prominent concomitantly with the maturation of osteocytes, chondrocytes, and skeletal muscle cells. RB1CC1 expression in these musculoskeletal cells increased with RB1 expression, which is linked to the terminal differentiation of many tissues and cells. In addition, the introduction of wild-type RB1CC1 decreased the formation of macroscopic colonies in the cell growth assay. Accordingly, both RB1CC1 and RB1 genes preferentially co-expressed and contributed to the maturation of human embryonic musculoskeletal cells, and may regulate the proliferative activity and maturation of tumor cells derived from these tissues.     

Rapid and efficient generation of lentivirally gene-modified dendritic cells from DC progenitors with bone marrow stromal cells

Since dendritic cells (DC) play pivotal roles in both innate and adaptive immunity, DC can be a good target for immuno-gene therapy. However, the optimal generation method for gene-modified DC has not yet been well exploited. CD34+ cells from cord blood (CB), bone marrow (BM), or peripheral blood (PB) were expanded in a medium containing stem cell factor (SCF), flt 3 ligand (Flt3L) and thrombopoietin (TPO) with or without HESS-5, a murine BM stromal cell line, for 2 weeks (the first expansion step), then differentiated to DC in a medium containing granulocyte-macrophage colony-stimulating factor (GM-CSF), flt 3 ligand (Flt3L), stem cell factor (SCF), tumor necrosis factor-alpha (TNF-alpha), IL-4, and lipopolysaccharide (LPS) for 9 days (the second differentiation step). DC progenitors were transduced with human immunodeficiency virus (HIV) vectors at different time points during the second step. Use of HESS-5 during the first step resulted in more DC generation than without it (cell expansion: CB, 10,461 vs. 354-fold; BM, 962 vs. 225-fold; peripheral blood mononuclear cell (PBMC), 8,506 vs. 240-fold; %DC: CB, 83.4% vs. 76.9%; BM, 83.6 vs. 69.8%; PBMC, 85.9 vs. 60.5%). Gene transduction to the in vitro expanded DC progenitors at day 3 during the second step, resulted in better final yield of the gene-modified DC than that to those at day 0 or day 6 (as much as 44% of DC expressed green fluorescence protein (GFP) as a transgene) and the transduction efficiency correlated with endocytic ability and percent of S phase. DC transduced with an HIV vector encoding a melanoma antigen, MART-1, were adequately recognized by specific anti-MART-1 CTL. The two-step culture method with HESS-5 is useful for rapid expansion of DC progenitors and subsequent lentiviral gene transduction to DC.     

Comparative genomic analysis of the MHC: the evolution of class I duplication blocks, diversity and complexity from shark to man

Summary: The major histocompatibility complex (MHC) genomic region is composed of a group of linked genes involved functionally with the adaptive and innate immune systems. The class I and class II genes are intrinsic features of the MHC and have been found in all the jawed vertebrates studied so far. The MHC genomic regions of the human and the chicken (B locus) have been fully sequenced and mapped, and the mouse MHC sequence is almost finished. Information on the MHC genomic structures (size, complexity, genic and intergenic composition and organization, gene order and number) of other vertebrates is largely limited or nonexistent. Therefore, we are mapping, sequencing and analyzing the MHC genomic regions of different human haplotypes and at least eight nonhuman species. Here, we review our progress with these sequences and compare the human MHC structure with that of the nonhuman primates (chimpanzee and rhesus macaque), other mammals (pigs, mice and rats) and nonmammalian vertebrates such as birds (chicken and quail), bony fish (medaka, pufferfish and zebrafish) and cartilaginous fish (nurse shark). This comparison reveals a complex MHC structure for mammals and a relatively simpler design for nonmammalian animals with a hypothetical prototypic structure for the shark. In the mammalian MHC, there are two to five different class I duplication blocks embedded within a framework of conserved nonclass I and/or nonclass II genes. With a few exceptions, the class I framework genes are absent from the MHC of birds, bony fish and sharks. Comparative genomics of the MHC reveal a highly plastic region with major structural differences between the mammalian and nonmammalian vertebrates. Additional genomic data are needed on animals of the reptilia, crocodilia and marsupial classes to find the origins of the class I framework genes and examples of structures that may be intermediate between the simple and complex MHC organizations of birds and mammals, respectively.     

Cytological features of dedifferentiated adenoid cystic carcinoma of the trachea

Adenoid cystic carcinoma (AdCC) is a distinct type of carcinoma, and cytological examination has been recognized as a useful tool in its diagnosis. Dedifferentiation is defined as the abrupt transformation of a low‐grade tumor into a tumor with high‐grade components. Albeit extremely rare, dedifferentiated AdCC has been reported: however, the cytological features of this tumor have not been documented. We observed a case in which a 66‐year‐old Japanese male had stenosis and thickness of the lower tracheal and bronchial walls. Cytological smears of a bronchial brush specimen revealed features typical for low‐grade AdCC. However, a few cohesive epithelial cell clusters composed of large, atypical polygonal cells with large nuclei and conspicuous nucleoli also were present. This component was considered to represent dedifferentiated carcinoma. Histopathological study of the resected bronchial tumor revealed dedifferentiated AdCC. The cytological diagnosis of conventional low‐grade AdCC is straightforward in most cases, although extremely rare, dedifferentiated carcinoma can occur within the conventional AdCC, and detection of a dedifferentiated component is possible in a cytological specimen because of obvious nuclear atypia. Therefore, careful observation is needed because cytologic diagnosis of dedifferentiated AdCC can help expedite treatment of this highly aggressive tumor. Diagn. Cytopathol. 2014;42:880–883. © 2013 Wiley Periodicals, Inc.     

Clear cell adenocarcinoma present exclusively within endometrial polyp: report of two cases

Endometrial polyp is a common benign lesion that protrudes into the endometrial surface. The incidence of carcinoma within endometrial polyp is thought to be low, however, postmenopausal women with endometrial polyps are at an increased risk. Endometrial clear cell adenocarcinoma is a distinct and relatively rare subtype of endometrial carcinoma, and recent studies have proposed putative precursor lesions of clear cell adenocarcinoma, namely clear cell endometrial glandular dysplasia (EmGD) and clear cell endometrial intraepithelial carcinoma (EIC). Herein, we describe two cases of clear cell adenocarcinoma present exclusively within endometrial polyp and discuss the association of its precursor. Two postmenopausal Japanese females, 66-year-old (Case 1) and 54-year-old (Case 2) presented with abnormal genital bleeding. Cytological examination of both cases revealed adenocarcinoma, thus, hysterectomy was performed. Histopathological studies demonstrated clear cell adenocarcinoma within exclusively endometrial polyp in both cases. The peculiar finding in Case 1 was presence of atypical glandular cells with large round to oval nuclei and clear cytoplasm within the atrophic endometrial glands in the surrounding endometrial tissue, which corresponded to clear cell EIC. A recent study showed that 33% of uteri had at least one focus of clear cell EmGD in endometrial polyps. Accordingly, clear cell adenocarcinoma and clear cell EmGD can occur in association with endometrial polyps more frequently than previously thought. Therefore, detailed histopathological examination is important in diagnosis of endometrial polyps, especially in the postmenopausal women, moreover cytological examination is a useful tool in the postmenopausal women with endometrial polyps.     

Clinicopathological features and CD57 expression in renal cell carcinoma in acquired cystic disease of the kidneys: with special emphasis on a relation to the duration of haemodialysis,the degree of calcium oxalate deposition,histological type,and possible tumorigenesis

Enoki Y, Katoh G, Okabe H & Yanagisawa A
(2010) Histopathology 56, 384–394 Clinicopathological features and CD57 expression in renal cell carcinoma in acquired cystic disease of the kidneys: with special emphasis on a relation to the duration of haemodialysis, the degree of calcium oxalate deposition, histological type, and possible tumorigenesis Aims: Acquired cystic disease of the kidney (ACDK) in patients undergoing haemodialysis is known to develop into renal cell carcinoma (RCC), but its pathogenesis remains unclear. The aims were to analyse the histological findings of ACDK‐RCC and to determine its histogenesis. Methods and results: Twenty‐nine RCCs in 23 patients with ACDK were classified into three groups according to the duration of haemodialysis and were analysed for histological type, calcium oxalate (Oxa) deposition, and cyst and atypical cyst (AC) formation. Histologically, 21 tumours were ACDK‐RCC and eight were clear cell carcinoma (CCC). The ratio of ACDK‐RCC and the numbers of cysts and ACs increased as the duration of haemodialysis was prolonged. The degrees of intratumoral Oxa deposition and cyst and AC formation of ACDK‐RCCs were higher than those of CCCs (Oxa, P = 0.028; cyst, P < 0.0001; AC, P = 0.0002). Many ACDK‐RCCs (85.7%) and some CCCs (50%) had characteristics of the thin ascending loop of Henle as assessed by CD57 (HNK‐1) expression, which was rarely expressed in the 29 control cases. Conclusions: ACDK‐RCCs reveal characteristics of Henle’s loop, which may be related to their peculiar pathological features, including intratumoral oxalate deposition and cyst and AC formation.