Autosomal recessive osteopetrosis (ARO) is a heterogeneous disorder, characterized by defective osteoclastic resorption of bone that results in increased bone density. We have studied nine individuals with an intermediate form of ARO, from the county of Västerbotten in Northern Sweden. All afflicted individuals had an onset in early infancy with optic atrophy, and in four patients anemia was present at diagnosis. Tonsillar herniation, foramen magnum stenosis, and severe osteomyelitis of the jaw were common clinical features. Whole exome sequencing, verified by Sanger sequencing, identified a splice site mutation c.212 + 1 G > T in the SNX10 gene encoding sorting nexin 10. Sequence analysis of the SNX10 transcript in patients revealed activation of a cryptic splice site in intron 4 resulting in a frame shift and a premature stop (p.S66Nfs * 15). Haplotype analysis showed that all cases originated from a single mutational event, and the age of the mutation was estimated to be approximately 950 years. Functional analysis of osteoclast progenitors isolated from peripheral blood of patients revealed that stimulation with receptor activator of nuclear factor kappa-B ligand (RANKL) resulted in a robust formation of large, multinucleated osteoclasts which generated sealing zones; however these osteoclasts exhibited defective ruffled borders and were unable to resorb bone in vitro.

Corneal dystrophies are a clinically and genetically heterogeneous group of inherited disorders that bilaterally affect corneal transparency. They are defined according to the corneal layer affected and by their genetic cause. In this study, we identified a dominantly inherited epithelial recurrent erosion dystrophy (ERED)-like disease that is common in northern Sweden. Whole-exome sequencing resulted in the identification of a novel mutation, c.2816C>T, p.T939I, in the COL17A1 gene, which encodes collagen type XVII alpha 1. The variant segregated with disease in a genealogically expanded pedigree dating back 200 years. We also investigated a unique COL17A1 synonymous variant, c.3156C>T, identified in a previously reported unrelated dominant ERED-like family linked to a locus on chromosome 10q23-q24 encompassing COL17A1. We show that this variant introduces a cryptic donor site resulting in aberrant pre-mRNA splicing and is highly likely to be pathogenic. Bi-allelic COL17A1 mutations have previously been associated with a recessive skin disorder, junctional epidermolysis bullosa, with recurrent corneal erosions being reported in some cases. Our findings implicate presumed gain-of-function COL17A1 mutations causing dominantly inherited ERED and improve understanding of the underlying pathology.

The importance of the tumour microbiome in different aspects of colorectal cancer (CRC) has been increasingly recognised, but many questions remain. The aim of this study was to explore the effect of specific CRC associated microbes on the tumour immune response, which has a considerable prognostic value in CRC. We applied specific qPCR to detect Parvimonas micra and Fusobacterium nucleatum in tumour tissues from an immunologically well-characterised cohort of 69 CRC patients. This cohort included detailed analyses of immune profiles based on flow cytometry and transcriptomics in tumour tissue and blood, along with comprehensive analyses of molecular subtypes. P. micra and F. nucleatum were detected in 24% and 64% of tumour tissues, respectively. We found a significant association of P. micra with high-grade tumours and tumours of CMS1 subtype. F. nucleatum was significantly associated with right-sided tumours, microsatellite instability, and CMS1 tumours. The immunological analyses revealed significant associations of P. micra with activated CD69+ T lymphocytes and increased antigen-presenting HLA-DR+ B lymphocytes. P. micra was also positively associated with M1 and M2 macrophage traits. The impact of P. micra tumour colonisation on the immune response was further assessed using transcriptomics in validation of our findings. No associations were found between F. nucleatum and immune profiles in this study. Our findings support novel associations between P. micra and the immune response in CRC. A better understanding of these interactions might help to identify important predictive and prognostic tools as well as new targets for therapy.

The local anti-tumour immune response has important prognostic value in colorectal cancer (CRC). In the era of immunotherapy, a better understanding of the immune response in molecular subgroups of CRC may lead to significant advances in personalised medicine. On this note, microsatellite instable (MSI) tumours have been characterised by increased immune infiltration, suggesting MSI as a marker for immune inhibitor checkpoint therapy. Here, we used flow cytometry to perform a comprehensive analysis of immune activity profiles in tumour tissues, adjacent non-malignant tissues and blood, from a cohort of 69 CRC patients. We found several signs of immune suppression in tumours compared to adjacent non-malignant tissues, including T cells more often expressing the immune checkpoint molecules programmed cell death protein (PD-1) and cytotoxic T lymphocyte-associated protein 4 (CTLA-4). We further analysed immune cell infiltration in molecular subgroups of CRC. MSI tumours were indeed found to be associated with increased immune infiltration, including increased fractions of PD-1+ T cells. No correlation was, however, found between MSI and the fraction of CTLA-4+ T cells. Interestingly, within the group of patients with microsatellite stable (MSS) tumours, some also presented with increased immune infiltration, including comparably high portions of PD-1+ T cells, but also CTLA-4+ T cells. Furthermore, no correlation was found between PD-1+ and CTLA-4+ T cells, suggesting that different tumours may, to some extent, be regulated by different immune checkpoints. We further evaluated the distribution of immune activity profiles in the consensus molecular subtypes of CRC. In conclusion, our findings suggest that different immune checkpoint inhibitors may be beneficial for selected CRC patients irrespective of MSI status. Improved predictive tools are required to identify these patients.

Gene co-expression networks (GCNs) can be prepared using a variety of mathematical approaches based on data sampled across diverse developmental processes, tissue types, pathologies, mutant backgrounds, and stress conditions. These networks are used to identify genes with similar expression dynamics but are prone to introducing false-positive and false-negative relationships, especially in the instance of large and heterogenous datasets. With the aim of optimizing the relevance of edges in GCNs and enhancing global biological insight, we propose a novel approach that involves a data-centering step performed simultaneously per gene and per sub-experiment, called centralization within sub-experiments (CSE). Using a gene set encoding the plant mitochondrial proteome as a case study, our results show that all CSE-based GCNs assessed had significantly more edges within the majority of the considered functional sub-networks, such as the mitochondrial electron transport chain and its complexes, than GCNs not using CSE; thus demonstrating that CSE-based GCNs are efficient at predicting canonical functions and associated pathways, here referred to as the core gene network. Furthermore, we show that correlation analyses using CSE-processed data can be used to fine-tune prediction of the function of uncharacterized genes; while its use in combination with analyses based on non-CSE data can augment conventional stress analyses with the innate connections underpinning the dynamic system being examined. Therefore, CSE is an effective alternative method to conventional batch correction approaches, particularly when dealing with large and heterogenous datasets. The method is easy to implement into a pre-existing GCN analysis pipeline and can provide enhanced biological relevance to conventional GCNs by allowing users to delineate a core gene network.