RESUMEN
The precise regulation of DNA replication is vital for cellular division and genomic integrity. Central to this process is the replication factor C (RFC) complex, encompassing five subunits, which loads proliferating cell nuclear antigen onto DNA to facilitate the recruitment of replication and repair proteins and enhance DNA polymerase processivity. While RFC1's role in cerebellar ataxia, neuropathy, and vestibular areflexia syndrome (CANVAS) is known, the contributions of RFC2-5 subunits on human Mendelian disorders is largely unexplored. Our research links bi-allelic variants in RFC4, encoding a core RFC complex subunit, to an undiagnosed disorder characterized by incoordination and muscle weakness, hearing impairment, and decreased body weight. We discovered across nine affected individuals rare, conserved, predicted pathogenic variants in RFC4, all likely to disrupt the C-terminal domain indispensable for RFC complex formation. Analysis of a previously determined cryo-EM structure of RFC bound to proliferating cell nuclear antigen suggested that the variants disrupt interactions within RFC4 and/or destabilize the RFC complex. Cellular studies using RFC4-deficient HeLa cells and primary fibroblasts demonstrated decreased RFC4 protein, compromised stability of the other RFC complex subunits, and perturbed RFC complex formation. Additionally, functional studies of the RFC4 variants affirmed diminished RFC complex formation, and cell cycle studies suggested perturbation of DNA replication and cell cycle progression. Our integrated approach of combining in silico, structural, cellular, and functional analyses establishes compelling evidence that bi-allelic loss-of-function RFC4 variants contribute to the pathogenesis of this multisystemic disorder. These insights broaden our understanding of the RFC complex and its role in human health and disease.
RESUMEN
Hermansky-Pudlak syndrome (HPS) is a group of rare genetic disorders, with several subtypes leading to fatal adult-onset pulmonary fibrosis (PF) and no effective treatment. Circulating biomarkers detecting early PF have not been identified. We investigated whether endocannabinoids could serve as blood biomarkers of PF in HPS. We measured endocannabinoids in the serum of HPS, IPF, and healthy human subjects and in a mouse model of HPSPF. Pulmonary function tests (PFT) were correlated with endocannabinoid measurements. In a pale ear mouse model of bleomycin-induced HPSPF, serum endocannabinoid levels were measured with and without treatment with zevaquenabant (MRI-1867), a peripheral CB1R and iNOS antagonist. In three separate cohorts, circulating anandamide levels were increased in HPS-1 patients with or without PF, compared to healthy volunteers. This increase was not observed in IPF patients or in HPS-3 patients, who do not have PF. Circulating anandamide (AEA) levels were negatively correlated with PFT. Furthermore, a longitudinal study over the course of 5-14 years with HPS-1 patients indicated that circulating AEA levels begin to increase with the fibrotic lung process even at the subclinical stages of HPSPF. In pale ear mice with bleomycin-induced HpsPF, serum AEA levels were significantly increased in the earliest stages of PF and remained elevated at a later fibrotic stage. Zevaquenabant treatment reduced the increased AEA levels and attenuated progression in bleomycin-induced HpsPF. Circulating AEA may be a prognostic blood biomarker for PF in HPS-1 patients. Further studies are indicated to evaluate endocannabinoids as potential surrogate biomarkers in progressive fibrotic lung diseases.
RESUMEN
OBJECTIVES: To identify the factors that can potentially affect the ability of electrodiagnostic tests such as sural/radial amplitude ratio or SRAR and the presence of carpal tunnel syndrome or CTS to detect early subclinical neuropathy in diabetes mellitus (DM). Likewise, to investigate the likelihood of developing subclinical neuropathy that can be detected by a positive CTS or SRAR abnormalities, because of the presence of anthropometric factors and sugar levels, in addition to DM duration METHODOLOGY: A retrospective cohort study was undertaken among 144 DM type 2 patients with confirmed subclinical neuropathy. Demographic data such as age, height and age, body mass index (BMI) and blood glucose profiles were obtained. Nerve conduction findings (SRAR and CTS protocols) were statistically analyzed using two sample t-test and multiple logistic regression ratios RESULTS: Patients who were positive in the CTS protocols were taller and had lower BMI. They had shorter duration of DM but with extreme elevations in blood glucose. Variables that are independently associated with a (+) CTS are duration of DM, FBS, BMI, height and weight. Patients with SRAR abnormalities were older and obese, with longer duration of DM and less marked elevations in blood glucose. Variables that are independently associated with SRAR abnormalities are age, duration of DM, weight and BMI CONCLUSION: Factors such as age, duration of DM, weight and height, BMI as well as glucose control can potentially affect the ability of various electrodiagnostic tests (SRAR and the presence of CTS) to detect early subclinical neuropathy. Since confounding factors was different between CTS and SRAR, the pathogenesis of these conditions may be different. The likelihood of developing subclinical neuropathy that can be detected by a (+) CTS or SRAR abnormalities because of the presence of certain factors, were documented.