Cytoskeleton stiffness regulates cellular senescence and innate immune response in Hutchinson-Gilford Progeria Syndrome.

Hutchinson-Gilford progeria syndrome (HGPS) is caused by the accumulation of mutant prelamin A (progerin) in the nuclear lamina, resulting in increased nuclear stiffness and abnormal nuclear architecture. Nuclear mechanics are tightly coupled to cytoskeletal mechanics via lamin A/C. However, the role of cytoskeletal/nuclear mechanical properties in mediating cellular senescence and the relationship between cytoskeletal stiffness, nuclear abnormalities, and senescent phenotypes remain largely unknown. Here, using muscle-derived mesenchymal stromal/stem cells (MSCs) from the Zmpste24-/- (Z24-/- ) mouse (a model for HGPS) and human HGPS fibroblasts, we investigated the mechanical mechanism of progerin-induced cellular senescence, involving the role and interaction of mechanical sensors RhoA and Sun1/2 in regulating F-actin cytoskeleton stiffness, nuclear blebbing, micronuclei formation, and the innate immune response. We observed that increased cytoskeletal stiffness and RhoA activation in progeria cells were directly coupled with increased nuclear blebbing, Sun2 expression, and micronuclei-induced cGAS-Sting activation, part of the innate immune response. Expression of constitutively active RhoA promoted, while the inhibition of RhoA/ROCK reduced cytoskeletal stiffness, Sun2 expression, the innate immune response, and cellular senescence. Silencing of Sun2 expression by siRNA also repressed RhoA activation, cytoskeletal stiffness and cellular senescence. Treatment of Zmpste24-/- mice with a RhoA inhibitor repressed cellular senescence and improved muscle regeneration. These results reveal novel mechanical roles and correlation of cytoskeletal/nuclear stiffness, RhoA, Sun2, and the innate immune response in promoting aging and cellular senescence in HGPS progeria.

Genomic instability and innate immune responses to self-DNA in progeria.

In the last decade, we have seen increasing evidence of the importance of structural nuclear proteins such as lamins in nuclear architecture and compartmentalization of genome function and in the maintenance of mechanical stability and genome integrity. With over 400 mutations identified in the LMNA gene (encoding for A-type lamins) associated with more than ten distinct degenerative disorders, the role of lamins as genome caretakers and the contribution of lamins dysfunction to disease are unarguable. However, the molecular mechanisms whereby lamins mutations cause pathologies remain less understood. Here, we review pathways and mechanisms recently identified as playing a role in the pathophysiology of laminopathies, with special emphasis in Hutchinson Gilford Progeria Syndrome (HGPS). This devastating incurable accelerated aging disease is caused by a silent mutation in the LMNA gene that generates a truncated lamin A protein "progerin" that exerts profound cellular toxicity and organismal decline. Patients usually die in their teens due to cardiovascular complications such as myocardial infarction or stroke. To date, there are no efficient therapies that ameliorate disease progression, stressing the need to understand molecularly disease mechanisms that can be targeted therapeutically. We will summarize data supporting that replication stress is a major cause of genomic instability in laminopathies, which contributes to the activation of innate immune responses to self-DNA that in turn accelerate the aging process.

Methionine restriction for improving progeria: another autophagy-inducing anti-aging strategy?

Methionine restriction, i.e., a partial depletion of the essential sulfur amino acid methionine from nutrition, extends lifespan in model organisms including yeast, nematodes, mice and rats. Recent results indicate that this strategy also prolongs health span and longevity in 2 short-lived strains of mice (with the LmnaG609G/G609G or zmpste24-/- genotypes) that represent animal models of Hutchinson-Gilford progeria syndrome (HGPS). The beneficial effects of methionine restriction on HGPS could be linked to reduced inflammation, and improved DNA stability, as well as the normalization of lipid and bile acid metabolism. Previous work has established that behavioral, nutritional, pharmacological and genetic manipulations that extend longevity in model organisms are only efficient if they induce increased autophagic flux. Methionine restriction extends lifespan in Saccharomyces cerevisiae in an Atg5- and Atg7-dependent fashion, supporting the notion that methionine restriction may indeed mediate its antiaging effects through the induction of macroautophagy/autophagy as well. Based on these findings, we speculate that autophagy might constitute an actionable therapeutic target to treat progeroid syndromes.

Insights into the role of immunosenescence during varicella zoster virus infection (shingles) in the aging cell model.

Varicella zoster virus (VZV) is the etiological agent of shingles, a painful skin rash that affects a significant proportion of the elderly population. In the present study, we used two aging cell models, Hutchinson-Gilford progeria syndrome (HGPS) fibroblasts and stress or replicative senescence-induced normal human dermal fibroblasts (NHDFs), to investigate age-associated susceptibility to VZV infection. VZV infectivity titers were significantly associated with donor age in HGPS fibroblasts and senescence induction in NHDFs. High throughput RNA-sequencing (RNA-seq) analysis was performed to assess global and dynamic changes in the host transcriptomes of VZV-infected aging cells. Analysis of differentially expressed genes (DEGs) indicated that VZV infection in aged HGPS fibroblasts resembled that in senescent NHDFs, particularly in terms of genes associated with pattern recognition receptors in virus sensing network, providing novel insights into the mechanisms of senescence-associated susceptibility to VZV infection. Additionally, we identified stimulator of interferon genes (STING) as a potential VZV sensing receptor. Knockdown of STING expression resulted in increased viral replication in primary fibroblasts, whereas STING overexpression led to suppression of VZV plaque formation. In conclusion, our findings highlight the important role of immunosenescence following VZV infection and provide significant insights into the mechanisms underlying cellular sensing of VZV infection and the induction of immune responses in aged skin cells.

Infection susceptibility and immune senescence with advancing age replicated in accelerated aging Lmna(Dhe) mice.

Aging confers increased susceptibility to common pathogens including influenza A virus. Despite shared vulnerability to infection with advancing age in humans and rodents, the relatively long time required for immune senescence to take hold practically restricts the use of naturally aged mice to investigate aging-induced immunological shifts. Here, we show accelerated aging Lmna(Dhe) mice with spontaneous mutation in the nuclear scaffolding protein, lamin A, replicate infection susceptibility, and substantial immune cell shifts that occur with advancing age. Naturally aged (≥ 20 month) and 2- to 3-month-old Lmna(Dhe) mice share near identically increased influenza A susceptibility compared with age-matched Lmna(WT) control mice. Increased mortality and higher viral burden after influenza infection in Lmna(Dhe) mice parallel reduced accumulation of lung alveolar macrophage cells, systemic expansion of immune suppressive Foxp3⁺ regulatory T cells, and skewed immune dominance among viral-specific CD8⁺T cells similar to the immunological phenotype of naturally aged mice. Thus, aging-induced infection susceptibility and immune senescence are replicated in accelerated aging Lmna(Dhe) mice.

Abnormalities of B cells and dendritic cells in SAMP1 mice.

The age-related changes in the function of antigen-presenting cells (APC) were examined using a substrain of senescence-accelerated mouse (SAMP1). In the primary mixed lymphocyte reaction (MLR), dendritic cells (DC) from aged SAMP1 mice showed less stimulatory activity than those of age-matched BALB/c or young SAMP1 mice. In the secondary MLR, the stimulatory activity of B cells was found to be lower in aged SAMP1 mice but not in age-matched BALB/c or young SAMP1 mice. In addition, these age-related decreases in the stimulatory activity of APC were found to be related to changes in the surface density of major histocompatibility complex class II and intercellular adhesion molecule-1 (ICAM-1) (but not B7-1 or B7-2 molecule) on APC (DC and B cells).

Exposure to low-frequency pulsed electromagnetic fields increases mitogen-induced lymphocyte proliferation in Down's syndrome.

We previously reported that exposure of human mitogen-stimulated peripheral blood lymphocytes (PBL) to extremely low frequency pulsed electromagnetic fields (PEMFs) could restore the defective proliferative capability of PBL from aged subjects. The effects of exposure to PEMFs were studied in PBL from 25 patients with Down's syndrome (DS), a syndrome of premature aging characterized by precocious immune system derangement, including age-related defective PBL proliferative capability. PBL were stimulated with different doses of phytohemagglutinin, and cell proliferation was assessed by measuring the incorporation of tritiated thymidine. After PEMF-exposure, a significant increase in cell proliferation was observed in cells from DS children and young adults, but it was much more evident in PBL from relatively aged DS patients. The age-related effect of PEMFs on DS lymphocytes demonstrates that age must be considered a major variable when studies on DS are performed, and confirms that DS must be regarded as a syndrome of accelerated aging.

Immunological aspects of progeria (Hutchinson-Gilford syndrome) in a 15-month-old child.

A thorough analysis of the immunological status was conducted in a 15-month-old child with progeria (Hutchinson-Gilford syndrome). Total leukocyte and neutrophil counts were slightly increased, and monocytes were decreased. Percentage and numbers of CD4+ cells in the blood were mildly decreased as well as the CD4/CD8 cell ratio. CD20 (B-cell marker) bearing cells and cells bearing Ia-antigens were increased, as well as CD16 and CD56 marker-bearing cells (natural-killer cells, NK). Lymphocyte proliferation upon stimulation with phytohaemagglutinin and purified protein derivative were decreased, and with pokeweed mitogen increased. NK cell activity appeared increased, particularly at lower effector: target cell ratios.

Hutchinson-Gilford progeria syndrome in a 45-year-old man.

A 45-year-old man with typical Hutchinson-Gilford progeria syndrome is described. The patient had the characteristic physical findings of this syndrome, such as short stature, "horse-riding" stance, coxa valga, alopecia, micrognathia, craniofacial disproportion, and prominent eyes. He had refractory congestive heart failure due to arteriosclerotic heart disease and hypertension, and he also had arteriosclerosis obliterans. Some immunologic and endocrinologic abnormalities commonly seen in the elderly were present in this patient. On the basis of a review of the literature, this is the first patient with this syndrome who had survived into the fourth decade.

Lymphocyte proliferation and nucleoid sedimentation in a case of premature aging distinct from Werner's syndrome.

Lymphocyte proliferation and nucleoid sedimentation were studied in a patient with premature aging resembling the Werner's syndrome (WS). Onset of patchy brown hyperpigmentations at the age of 9 months permitted distinction from classical WS and suggested a WS-like premature aging disease. By photometric recording of density changes during cell culture, we examined the course of cell proliferation after PHA stimulation over 7 days and compared these results to those obtained in two normal controls. Cultured cells of the patient displayed an aberrant proliferation pattern characterized by continuous growth without an initial reduction phase. The markedly reduced proliferative capacity of purified cells from the patient could in part be corrected by fetal bovine serum. The cells of the patient displayed a characteristic nucleoid sedimentation profile after ultraviolet irradiation indicating retarded DNA replication, which may be a common feature of various premature aging diseases. The absence of thermolability of cell proliferation and the presence of a high number of chromatid aberrations disclosed differences from classical WS.

Subpopulations of human T lymphocytes. XVII. Imbalance of T cell subsets in patients with progeria and Werner's syndrome.

Peripheral blood from 4 patients with progeria and their family members and 2 patients with Werner's syndrome were analyzed for the proportion of total T cells and T cells with receptors for IgM (Tmu) or IgG (Tgamma). In 3 patients proportions of Tmu cells were within the range for age- and sex-matched controls. 1 patient had significantly lower proportions of Tmu cells when compared to age- and sex-matched controls. The proportions of Tgamma cells were increased in 2 and were within the range for normal controls in the remaining 2 patients. When data were analyzed with regard to Tmu/Tgamma ratios, 3 of 4 patients exhibited ratios lower than the lowest limit observed in age- and sex-matched controls. Surprisingly, family members in all four families exhibited patterns of Tmu and Tgamma cells similar to that seen in the patient. Both patients with Werner's syndrome had proportions of Tgamma cells, resulting in low Tmu/Tgamma ratios.

Detection of HLA antigens on progeria syndrome fibroblasts.

It has been suggested that cultured skin fibroblasts derived from patients with progeria, a premature aging syndrome, have absent or markedly reduced HLA antigen expression, thus implicating a possible immunological role in the pathogenesis of this disease. An analysis of HLA expression in nine progeric fibroblast strains and related familial strains is reported. Normal HLA expression and inheritance were found. No significant association between HLA type and progeria was detected. Therefore, we conclude an abnormality of HLA expression is unlikely to be an underlying defect in progeria.

T cells, precocious aging, and familial neoplasia.

A 15-year-old girl presented with precocious aging and was found to have low levels of active and total T cells. Family history revealed a high familial incidence of cancer on both the maternal and paternal sides, and activ T cell levels were found to be low in several living family members. The patient developed osteogenic sarcoma 13 months after initial study. Since our previous studies have reported low active and total T cells in patients with cancer, the present results suggest that subjects with low active T cells should be monitored frequently to detect possible neoplasia in it early stages. They also suggest that impaired cellular immunity in humans is associated with, if not the cause of, accelerated aging.

[Immunocytogenesis and formation of antibodies in experimentally aged animals (author's transl)]. / Uber die Immunozytogenese und Synthese der Antikörper bei experimentell gealterten Ratten

The authors use white rats with experimentally induced Progeria-like syndrome of Selye. The immunologic response following inoculation with the extracellular microbe-parasite D, pneumoniae and the intracellular parasite Br. abortus 19 is studied serologically, cellularly and biochemically. The experimentally aged rats show diminished and retarded plasmocyte reaction and weaker and slower agglutinine synthesis as compared with the controls. The immunochemical analysis of the serums of the experimentally aged rats reveals changes in the serum fractions and an affected ratio of albumin/globulin. The data suggest a disturbed adaptation and a reversal to an older ontogenic form of the aged organism.

[Cellular immunity against bacteria (intracellular and extracellular parasites) in experimentally aged rats (author's transl)]. / Zelluläre Immunität gegunüber Bakterien (intra- und extrazelluläre Parasiten) bei experimentell gealterten Ratten

The authors study the activity of RES in rats with Progeria-like syndrome of Selye, at the occasion of a repeated infection with bacteria--intracellular parasites (Brucella abortus 19) and bacteria--extracellular parasites (Diplococcus pneumoniae). They establish that immunization improves the activity of RES of the experimentally aged rats; still, its phagocytic and digestive functions remain by far feebler than those of the rats unsubjected to experimental ageing. By the aged animals the RES insufficiency is more pronounced towards the intracellular bacterium Brucella abortus 19, than towards the extracellular Diplococcus pneumoniae.

[Resistance against intra- and extracellular bacteria in experimentally aged rats (author's transl)]. / Resistenz gegen intra- und extrazelluläre Bakterien bei experimentell gealterten Ratten

The authors study the cellular activity of RES against the microbes intracellular and extracellular parasites in rats with pronounced Progeria-like syndrome of Selye. RES of the experimentally aged animals copes, through with a certain delay, with the invading extracellular bacterium Diplococcus pneumoniae, but is utterly helpless against the intracellular microbe Br. abortus 19. These relations are completly inverted in the rats that have not been experimentally aged.

[Morphology of the cells in the peritoneal exsudate and phosphatase activity of the peritoneal macrophages experimentally aged rats treated with the intracellular bacterium Brucella abortus 19 (author's transl)]. / Morphologie der Zellen im peritonealen Exsudat und die Phosphatase-Aktivität der peritonealen Makrophagen von experimentell gealterten Ratten, behandelt mit Brucella abortus 19

The authors study the morphology of the cells in the peritoneal exsudate and the phosphatase activity of the peritoneal macrophages, obtained from rats with Selye's Progeria-like syndrome. Judging by their morphologic characteristics the macrophages of the experimentally aged rats, before and after contact with Brucella abortus 19 do not differ from those obtained from the unsubjected to experimental ageing rats. The acid phosphatase activity and the adenosinetriphosphatase activity of the macrophages prior to contact with Brucella abortus 19 are nearly identical for both groups of animals. Following contact the activity of the enzymes increases but this increase is slower by aged rats and reaches its maximum 1--2 days later.