Cancer stem cells (CSCs) or tumor-initiating cells (TICs) are thought to be the main drivers for disease progression and treatment resistance across various cancer types. Identifying and targeting ...these rare cancer cells, however, remains challenging with respect to therapeutic benefit. Here, we report the enrichment of LGR5 expressing cells, a well-recognized stem cell marker, in mouse liver tumors, and the upregulation of LGR5 expression in human hepatocellular carcinoma. Isolated LGR5 expressing cells from mouse liver tumors are superior in initiating organoids and forming tumors upon engraftment, featuring candidate TICs. These cells are resistant to conventional treatment including sorafenib and 5-FU. Importantly, LGR5 lineage ablation significantly inhibits organoid initiation and tumor growth. The combination of LGR5 ablation with 5-FU, but not sorafenib, further augments the therapeutic efficacy in vivo. Thus, we have identified the LGR5
compartment as an important TIC population, representing a viable therapeutic target for combating liver cancer.
Megacystis microcolon intestinal hypoperistalsis syndrome (MMIHS) is a congenital visceral myopathy characterized by severe dilation of the urinary bladder and defective intestinal motility. The ...genetic basis of MMIHS has been ascribed to spontaneous and autosomal dominant mutations in actin gamma 2 (ACTG2), a smooth muscle contractile gene. However, evidence suggesting a recessive origin of the disease also exists. Using combined homozygosity mapping and whole exome sequencing, a genetically isolated family was found to carry a premature termination codon in Leiomodin1 (LMOD1), a gene preferentially expressed in vascular and visceral smooth muscle cells. Parents heterozygous for the mutation exhibited no abnormalities, but a child homozygous for the premature termination codon displayed symptoms consistent with MMIHS. We used CRISPR-Cas9 (CRISPR-associated protein) genome editing of Lmod1 to generate a similar premature termination codon. Mice homozygous for the mutation showed loss of LMOD1 protein and pathology consistent with MMIHS, including late gestation expansion of the bladder, hydronephrosis, and rapid demise after parturition. Loss of LMOD1 resulted in a reduction of filamentous actin, elongated cytoskeletal dense bodies, and impaired intestinal smooth muscle contractility. These results define LMOD1 as a disease gene for MMIHS and suggest its role in establishing normal smooth muscle cytoskeletal–contractile coupling.
Megacystis microcolon intestinal hypoperistalsis syndrome (MMIHS) is a congenital disorder characterized by loss of smooth muscle contraction in the bladder and intestine. To date, three genes are ...known to be involved in MMIHS pathogenesis: ACTG2, MYH11, and LMOD1. However, for approximately 10% of affected individuals, the genetic cause of the disease is unknown, suggesting that other loci are most likely involved. Here, we report on three MMIHS-affected subjects from two consanguineous families with no variants in the known MMIHS-associated genes. By performing homozygosity mapping and whole-exome sequencing, we found homozygous variants in myosin light chain kinase (MYLK) in both families. We identified a 7 bp duplication (c.3838_3844dupGAAAGCG p.Glu1282_Glyfs∗51) in one family and a putative splice-site variant (c.3985+5C>A) in the other. Expression studies and splicing assays indicated that both variants affect normal MYLK expression. Because MYLK encodes an important kinase required for myosin activation and subsequent interaction with actin filaments, it is likely that in its absence, contraction of smooth muscle cells is impaired. The existence of a conditional-Mylk-knockout mouse model with severe gut dysmotility and abnormal function of the bladder supports the involvement of this gene in MMIHS pathogenesis. In aggregate, our findings implicate MYLK as a gene involved in the recessive form of MMIHS, confirming that this disease of the visceral organs is heterogeneous with a myopathic origin.
We have succeeded in establishing 91 organoid lines from 128 primary mouse liver tumors. These organoids can be grown in long-term cultures, expand in vitro and initiate tumor in vivo.
Abstract
The ...current understanding of cancer biology and development of effective treatments for cancer remain far from satisfactory. This in turn heavily relies on the availability of easy and robust model systems that resemble the architecture/physiology of the tumors in patients to facilitate research. Cancer research in vitro has mainly been based on the use of immortalized 2D cancer cell lines that deviate in many aspects from the original primary tumors. The recent development of the organoid technology allowing generation of organ-buds in 3D culture from adult stem cells has endowed the possibility of establishing stable culture from primary tumors. Although culturing organoids from liver tumors is thought to be difficult, we now convincingly demonstrate the establishment of organoids from mouse primary liver tumors. We have succeeded in culturing 91 lines from 129 liver tissue/tumors. These organoids can be grown in long-term cultures in vitro. About 20% of these organoids form tumors in immunodeficient mice upon (serial) transplantation, confirming their tumorigenic and self-renewal properties. Interestingly, single cells from the tumor organoids have high efficiency of organoid initiation, and a single organoid derived from a cancer cell is able to initiate a tumor in mice, indicating the enrichment of tumor-initiating cells in the tumor organoids. Furthermore, these organoids recapitulate, to some extent, the heterogeneity of liver cancer in patients, with respect to phenotype, cancer cell composition and treatment response. These model systems shall provide enormous opportunities to advance our research on liver cancer (stem cell) biology, drug development and personalized medicine.
Mammoth Mountain, California, is a dacitic volcano that has experienced several periods of unrest since 1989. The onset of diffuse soil CO2 emissions at numerous locations on the flanks of the ...volcano began in 1989–1990 following an 11-month period of heightened seismicity. CO2 emission rates were measured yearly from 1995 to 2013 at Horseshoe Lake (HSL), the largest tree kill area on Mammoth Mountain, and measured intermittently at four smaller degassing areas around Mammoth from 2006 to 2013. The long-term record at HSL shows decadal-scale variations in CO2 emissions with two peaks in 2000–2001 and 2011–2012, both of which follow peaks in seismicity by 2–3years. Between 2000 and 2004 emissions gradually declined during a seismically quiet period, and from 2004 to 2009 were steady at ~100metric tonnes per day (td−1). CO2 emissions at the four smaller tree-kill areas also increased by factors of 2–3 between 2006 and 2011–2012, demonstrating a mountain-wide increase in degassing. Delays between the peaks in seismicity and degassing have been observed at other volcanic and hydrothermal areas worldwide, and are thought to result from an injection of deep CO2-rich fluid into shallow subsurface reservoirs causing a pressurization event with a delayed transport to the surface. Such processes are consistent with previous studies at Mammoth, and here we highlight (1) the mountain-wide response, (2) the characteristic delay of 2–3years, and (3) the roughly decadal reoccurrence interval for such behavior. Our best estimate of total CO2 degassing from Mammoth Mountain was 416td−1 in 2011 during the peak of emissions, over half of which was emitted from HSL. The cumulative release of CO2 between 1995 and 2013 from diffuse emissions is estimated to be ~2–3Mt, and extrapolation back to 1989 gives ~4.8Mt. This amount of CO2 release is similar to that produced by the mid-sized (VEI 3) 2009 eruption of Redoubt Volcano in Alaska (~2.3Mt over 11months), and significantly lower than long-term emissions from hydrothermal areas such as Solfatara in Campi Flegrei, Italy (16Mt over 28years).
•CO2 emissions and shallow seismicity are correlated at Mammoth Mtn. and vary on decadal timescales.•Increased CO2 emissions from Mammoth Mtn. lag periods of increased shallow seismicity by 2–3years.•Long-term regional measurements show a mountain-wide response to increased degassing.•Total CO2 from diffuse degassing is estimated to be ~2–3Mt for 1995–2013, and ~4.8Mt back to 1989.
Autoimmune hepatitis (AIH) develops in genetically predisposed individuals after an inciting or environmental trigger. These factors are unknown but may include viral infections, environmental ...toxins, drugs and vaccinations. Few reports are written about vaccination as potential trigger of autoimmune hepatitis. In this article, we additionally describe two vaccine-related cases of AIH. In both cases, long-term immune-suppressive therapy is demanded. Moreover, we present the cases of vaccine-related AIH from literature and compare these with idiopathic AIH and our own cases.
The 2009 eruption of Redoubt Volcano, Alaska, provided a rare opportunity to compare satellite measurements of sulfur dioxide (SO2) by the Ozone Monitoring Instrument (OMI) with airborne SO2 ...measurements by the Alaska Volcano Observatory (AVO). Herein we: (1) compare OMI and airborne SO2 column density values for Redoubt's tropospheric plume, (2) calculate daily SO2 masses from Mount Redoubt for the first three months of the eruption, (3) develop simple methods to convert daily measured SO2 masses into emission rates to allow satellite data to be directly integrated with the airborne SO2 emissions dataset, (4) calculate cumulative SO2 emissions from the eruption, and (5) evaluate OMI as a monitoring tool for high-latitude degassing volcanoes. A linear correlation (R2~0.75) is observed between OMI and airborne SO2 column densities. OMI daily SO2 masses for the sample period ranged from ~60.1kt on 24 March to below detection limit, with an average daily SO2 mass of ~6.7kt. The highest SO2 emissions were observed during the initial part of the explosive phase and the emissions exhibited an overall decreasing trend with time. OMI SO2 emission rates were derived using three methods and compared to airborne measurements. This comparison yields a linear correlation (R2~0.82) with OMI-derived emission rates consistently lower than airborne measurements. The comparison results suggest that OMI's detection limit for high latitude, springtime conditions varies from ~2000 to 4000t/d. Cumulative SO2 masses calculated from daily OMI data for the sample period are estimated to range from 542 to 615kt, with approximately half of this SO2 produced during the explosive phase of the eruption. These cumulative masses are similar in magnitude to those estimated for the 1989–90 Redoubt eruption. Strong correlations between daily OMI SO2 mass and both tephra mass and acoustic energy during the explosive phase of the eruption suggest that OMI data may be used to infer relative eruption size and explosivity. Further, when used in conjunction with complementary datasets, OMI daily SO2 masses may be used to help distinguish explosive from effusive activity and identify changes in lava extrusion rates. The results of this study suggest that OMI is a useful volcano monitoring tool to complement airborne measurements, capture explosive SO2 emissions, and provide high temporal resolution SO2 emissions data that can be used with interdisciplinary datasets to illuminate volcanic processes.
► Linear correlations between OMI and airborne SO2 column densities are found. ► A linear correlation between OMI-derived and airborne SO2 emission rates is found. ► OMI-derived emission rates underestimate SO2 relative to airborne measurements. ► Correlations between daily SO2 masses and eruptive activity are observed. ► OMI is a useful tool for measuring volcanic SO2 emissions remotely.
No single reliable parameter exists to assess liver graft function of extended criteria donors during ex-vivo normothermic machine perfusion (NMP). The liver maximum capacity (LiMAx) test is a ...clinically validated cytochromal breath test, measuring liver function based on 13CO2 production. As an innovative concept, we aimed to integrate the LiMAx breath test with NMP to assess organ function. Eleven human livers were perfused using NMP. After one hour of stabilization, LiMAx testing was performed. Injury markers (ALT, AST, miR-122, FMN, and Suzuki-score) and lactate clearance were measured and related to LiMAx values. LiMAx values ranged between 111 and 1838 µg/kg/h, and performing consecutive LiMAx tests during longer NMP was feasible. No correlation was found between LiMAx value and miR-122 and FMN levels in the perfusate. However, a significant inverse correlation was found between LiMAx value and histological injury (Suzuki-score, R = - 0.874, P < 0.001), AST (R = - 0.812, P = 0.004) and ALT (R = - 0.687, P = 0.028). Furthermore, a significant correlation was found with lactate clearance (R = 0.683, P = 0.043). We demonstrate, as proof of principle, that liver function during NMP can be quantified using the LiMAx test, illustrating a positive correlation with traditional injury markers. This new breath-test application separates livers with adequate cytochromal liver function from inadequate ones and may support decision-making in the safe utilization of extended criteria donor grafts.
Aberrant DNA methylation changes have been reported to be associated with carcinogenesis in cirrhotic HCC, but DNA methylation patterns for these non-cirrhotic HCC cases were not examined. Therefore, ...we sought to investigate DNA methylation changes on non-cirrhotic HCC using reported promising DNA methylation markers (DMMs), including HOXA1, CLEC11A, AK055957, and TSPYL5, on 146 liver tissues using quantitative methylation-specific PCR and methylated DNA sequencing. We observed a high frequency of aberrant methylation changes in the four DMMs through both techniques in non-cirrhotic HCC compared to cirrhosis, hepatitis, and benign lesions (p < 0.05), suggesting that hypermethylation of these DMMs is specific to non-cirrhotic HCC development. Also, the combination of the four DMMs exhibited 78% sensitivity at 80% specificity with an AUC of 0.85 in discriminating non-cirrhotic HCC from hepatitis and benign lesions. In addition, HOXA1 showed a higher aberrant methylation percentage in non-cirrhotic HCC compared to cirrhotic HCC (43.3% versus 13.3%, p = 0.039), which was confirmed using multivariate linear regression (p < 0.05). In summary, we identified aberrant hypermethylation changes in HOXA1, CLEC11A, AK055957, and TSPYL5 in non-cirrhotic HCC tissues compared to cirrhosis, hepatitis, and benign lesions, providing information that could be used as potentially detectable biomarkers for these unusual HCC cases in clinical practice.
Hepatic angiomyolipoma (HAML) may easily be misdiagnosed as a malignancy. The study aim was to assess diagnostic dilemmas, clinical management and outcome of this rare tumor.
This retrospective ...international multicenter study included all patients with pathologically proven HAML diagnosed between 1997 and 2017. Data on patient characteristics, diagnostic work-up, management and follow-up were analyzed.
Thirty-eight patients were included, 32 female. Median age was 56yrs (i.q.r. 43–64) and median HAML-diameter was 57.5 mm (i.q.r. 38.5–95.3). Thirty patients had undergone CT and 27/38 MRI of the liver, diagnostic biopsy was performed in 19/38. Initial diagnosis was incorrect in 15/38 patients, of which 13 were thought to have malignancy. In 84% biopsy resulted in a correct preoperative diagnosis. Twenty-nine patients were managed with surgical resection, 4/38 with surveillance and 3/38 with liver transplantation. Recurrence after resection occurred in two cases. No HAML related deaths or progression to malignancy were documented.
HAML diagnosis proved problematic even in hepatobiliary expertise centers. Biopsy is indicated and may provide valuable additional information when HAML diagnosis is considered on cross-sectional imaging, especially when surgical resection imposes a risk of complications. Conservative management with regular imaging follow-up might be justified when biopsy confirms (classic type) HAML.
PDF
