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Age-related thymic involution is characterized by a progressive regression in thymus size and a diminishment of thymic structure. A decrease in thymic compartments leads to the reduction of thymopoiesis. Thymic involution is closely associated with immunosenescence, a degeneration of the immune system primarily due to the alterations in T-cell composition. Strategies to improve the consequences of the aging thymus are currently under investigation. A wide array of knowledge has revealed a series of factors that are essential in the overall determination of thymic function and immune response. Evidence indicates that early programming of the thymus, sexual dimorphism, and the efficiency of specific T-cell progenitors and the thymic microenvironment are all crucial determinants of immune activity from early life through advanced ages. To fully understand the processes involved in age-related thymic involution, such determinants must be considered. The central purpose of this review is to emphasize previous and most recent evidence suggesting that these factors contribute to the influence of long-term immunity and ultimately shape the progression of thymic involution in advanced age.
As living standards and overall healthcare improve worldwide, the global population steps into a new era composed of a greater proportion of individuals of advanced ages. This proportion will continue to increase, as it is predicted that people approximately 60 years or older are estimated to account for 25 % of the total U.S. population by the year 2050. Despite such beneficial outcomes in terms of life expectancy, the expansion of aging populations induces undesirable stresses on public healthcare since prolonged life also carries negative consequences. The natural process of aging is concomitant with immunosenescence, a degeneration of the immune system that is characteristic of a greater susceptibility to infections [1, 2], an inadequate immune response to vaccinations [3–5] and an increased propensity for autoimmune diseases and cancers [6–8]. In fact, according to the U.S. Center for Disease Control (CDC), approximately 80 % of aged individuals are afflicted with at least one chronic disease as a result of a declination of immune function.
The prevalence of these age-related conditions is believed to ascribe primarily to qualitative and quantitative alterations of T-cell. In the elderly, naive T-cell production declines with an increased proportion of memory T-cell with oligoclonal expansion . This results in shrinkage of the T-cell receptor (TCR) repertoire and impaired T-cell functions [10, 11]. T cells of aged subjects phenotypically express more senescence markers such as CD57, and less homing receptors such as CD62L and CCR7, which are essential for their mobilization to proper site of action. In those of advanced ages, T cells undergo replicative senescence as attributed to the corrosion of telomeres, exhibit cell cycle arrest, and have limited proliferation in response to antigen stimulation . Aging is also manifested in the fact that naive T cells are generated in an extremely slow pace after allogeneic stem cell transplantation [12, 13].
The thymus is the primary organ responsible for de novo generation of immunocompetent T cells with a diverse repertoire of antigen-recognition. However, it is the organ being most ostensibly seen to decrease in size along with age. This process, termed as age-related thymic involution, is evolutionarily and conservatively maintained in vertebrates . The main components of the thymus include thymocytes of hematopoietic origin and thymic epithelial cells (TECs) of non-hematopoietic origin. The latter comprises the main stromal niche, termed as thymic epithelial space, for supporting T-cell development and maturation. The thymic epithelium starts to decrease as a main feature of age-related thymic involution from as early as the first year of human life at a rate of 3 % per year during adulthood, paralleling with an expansion of perivascular space that progressively fills with adipocytes and peripheral lymphocytes [15, 16]. Loss of demarcation of the thymic medulla and cortex and a disorganization of the corticomedullary junction were observed in the aged thymus [15–17]. Age-related thymic involution results in the reduction of thymopoiesis which precedes T-cell related immuno-incompetence in an advanced age.
Despite significant thymic atrophy, thymopoiesis does not cease completely in advanced age . Strategies to improve thymopoiesis in the aging thymus are actively under investigation [19–21]. To develop proper therapeutics, one must first consider the elements involved in the regulation of thymic involution overtime. Evidence indicates that early programming of the thymus is crucial in the determination of long-term immunological effects in adulthood. Additionally, sex differences in thymus size, thymopoiesis and involution play roles in deciding the level of immune function. Lastly, the crosstalk between T-cell progenitors and the thymic stroma is also critical in the influence of thymic function along with progressive aging. The purpose of this review is to summarize recent findings and current knowledge to understand how these elements shape the thymic size, involution and thymopoiesis.
Initial stocks of T cells with a variety of TCR repertoire are built during fetal and early post-natal life depending on thymus function. Early life programming of the thymus is crucial, having long-term effects on T-cell development. There is a great deal of evidence to support that programming of the thymus is shaped by both genetic and environmental factors.
Genetics plays a significant role in determining initial thymic size and rate of involution as has been shown in different strains of mice [22–24]. Studies on C57BL/6J_DBA/2J recombinant-inbred mice revealed that genes on chromosomes 9 and 10 determine initial thymus size . By comparing slow involution and fast involution BXD recombinant-inbred strains, Wang and colleagues found that fast involution strains exhibit a block in thymocyte development and a decrease in thymopoiesis . Genetic programming of initial thymus size and thymic involution rate may partially explain why there is a differential susceptibility to various pathogen invasions and spontaneous tumors among different mouse strains. The importance of initial thymus size could further be understood from recent studies on MCL1 transgenic mice. The female MCL1 transgenic mouse was found to have an enlarged thymus starting from at least 1 month of age. Despite age-related thymic involution, thymic enlargement is sustained in MCL1 mice at 7 months of age in relation to age- and sex-matched controls, suggesting that an initially enlarged thymus results in an attenuated thymic involution. In addition, the initially enlarged thymus established a higher steady-state capacity of thymopoiesis, a beneficial effect that could be seen when mice receive γ irradiation. After whole body γ irradiation, transgenic mice maintained more radioresistant resident primitive CD4-CD8- double negative 1 (DN1) cells, which reconstituted the thymus to a size larger than control after 7 and 15 days . Specific genetic disorders, such as DiGeorge syndrome and Down’s syndrome, also result in a series of immune deficiencies by affecting thymus development . In the case of DiGeorge syndrome, patients have little to no thymus and present a reduction of naive T cells and an oligoclonal expansion of T-cell repertoire, increasing their vulnerability to pathogenic infections and blood malignancies that are similarly noted in those of advanced ages [27, 28]. Accelerated immunoscenescence under these conditions recapitulates the fact that thymus function modified by early events has long-term effects on adult immunity.
In addition to genetics, environmental influences during early life have also been shown to control initial thymus size and long-term effects on cell-mediated immunity throughout adulthood and advanced age . Zinc deficiency, for example, results in thymic atrophy in malnourished children who also show an increased susceptibility to infections . In individuals of advanced age, the reduction of the zinc level in the body directly correlates with thymic involution and consequent immunological dysfunction . A restoration of microarchitecture and weight in the atrophic thymus was achieved in response to a 3–6 month in vivo supplementation of zinc . Furthermore, a longitudinal study conducted in Filipino adolescents revealed that prenatal malnutrition, the duration of exclusive breastfeeding and length increment during the first year of life, are all significantly associated with thymic function . An additional study in Bangladeshi infants demonstrates that thymic volume has a strong correlation with birth weight and duration of exclusive breastfeeding . In mice, a recent study monitoring postnatal thymus growth in males from 2 to 12 weeks of age demonstrated that maternal protein restriction influences thymic growth in early adult life. Interestingly, this study illustrated that maternal protein restriction during pregnancy has different effects on thymocyte proliferation and thymus growth by 12 weeks of age in comparison to protein restriction during lactation . This exemplifies that the thymic development is very sensitive to early events during both prenatal and early postnatal periods. Thus, in the long run, various factors involved in the regulation of early life programming have a long-term determination of the effects manifested in adulthood.
The importance of fetal and early infant programming of thymus function is highlighted by the acceleration of T-cell aging after thymectomy in infants and children with congenital heart defects (CHDs). The thymus is removed during the surgical correction of CHDs due to its anatomical obstruction in relation to the heart. A wide array of data shows that thymectomy during infancy results in a decreased number of CD4 and CD8 T cells, a diminished TCR repertoire and thymopoiesis, an increased amount of plasma IL-7, and a disequilibrium of the ratio of naive to memory T cells, all of which mimic changes expected to occur after physiological thymic involution in the elderly . Interestingly, a study in which T-cell composition was monitored for 3 decades in infants who had thymectomies shows that the removal of the thymus has a deleterious impact on T-cell for only the first 5 years after surgery. After this time frame, naive T-cell counts and signal joint TCR excision circles (sjTRECs), an indicator of recent thymic emigrants, return to values within the normal range. This observation could be explained by the functional regrowth of residue thymic tissue left behind during surgery . Since the practice of thymectomy during cardiac surgery only began 30–40 years ago and patients who underwent this surgery are currently young, further investigation is needed to determine whether thymectomy has an impact on T-cell development during advanced age. Studies revealed that the serological positive of cytomegalovirus (CMV) is closely associated with the severity of the impact on adult immune system after thymectomy in early life . Given the ubiquitous infection of CMV in human, the risk of early life thymectomy should be fully reevaluated.
For quite some time, differences in immune function between females and males have been well observed. Women generally have a lesser incidence and severity of viral, bacterial, fungal, and other parasitic diseases in relation to men [39, 40]. Women have a higher risk of a variety of autoimmune diseases such as multiple sclerosis, rheumatoid arthritis, and lupus . Genetics is a major determinant of immune function underlined by the fact that many genes implicated in critical immune function are located on the X-chromosome. Genes coding for androgen receptors, common gamma chains involved in the reception of signals from a variety of cytokines, and FOXP3 involved in regulatory T-cell development are all located on chromosome X. Dosage compensation for inactivation of X-linked genes in females serves as a genetic predisposition for differential immunological outcomes . Other than genetic disparities, hormonal level and receptor distribution are also believed to modulate the different immune responses observed between the sexes. The effect of sex hormones on the immune response can be exemplified by changes in Th1 and Th2 cytokine profiles during menopause and pregnancy when estrogen and progesterone levels fluctuate, in addition to the prominent observation that disease activities of multiple sclerosis and rheumatoid arthritis decrease dramatically during pregnancy [43, 44].
Thymic size is influenced by a series of hormones, including sex steroids and those involved in the hypothalamic-pituitary-adrenal axes . It has been revealed that both thymocytes and TECs express sex hormones receptors [45, 46]. Castration of male rodents results in significant enlargement of thymus [46, 47]. In addition, the size of the thymus has also been noted to fluctuate during pregnancy . Thymic involution is notably observed around the time of puberty when sex steroid production increases. To date, there is little literature that documents differences in thymic involution between males and females. The reason for this is two-fold; for one, sexual dimorphism may be overlooked by researchers focusing solely on the shrinkage of thymic size during aging in both sexes, not comparing differences in involution rates between females and males. Also, it is logical for investigators to design their experiments with mice that are of a particular sex to simplify data interpretation with fewer variables given that age differences already contribute to the variability. Nevertheless, there have been some analyses noting age-related involution differences in sexual dimorphism studies. Studies have shown that right before a decrease in thymic weight around the time of puberty, there is a short period where the thymic size decreases and then grows back. A decrease in thymic weight is notable in mice during days 18 to 26 after birth. Interestingly, thymic weight begins to steadily increase after day 26 in both sexes but lasts for a longer period of time in females (day 46) than in males (day 35). After this, an appreciable, one-way thymic involution begins around puberty with a more prominent declination observed in males than in females. It has been reported that there are twice as many thymocytes present in the female thymus after 9 months of age in comparison to age-matched males . A recent experiment conducted to compare thymic cellularity between age-matched C57/BL6 mice revealed a constant rate of involution in females but a biphasic speed of involution in males in mice 1–7 months of age. In males, a precipitous drop in thymic cellularity occurs from 1 to 3 months of age followed by a lower slope of involution from 3 to 7 months old. The greatest difference in thymic cellularity between the sexes that occurs during involution is most notable around 3 months of age (Fig. 1). Sexual dimorphism is also maintained in a MCL1 transgenic animal model. From 1 to 3 months old, transgenic MCL1 male mice exhibit a greater reduction in thymic cellularity than transgenic MCL1 females (Fig. 1). Unexpectedly, the presentation of the gene function of MCL1 is different between the sexes. Female MCL1 transgenic mouse but not male transgenic mouse was found to have an enlarged thymus. It was suggested that an in vivo environment in males suppresses the effect of MCL1 transgene since both female and male mice express same dose of transgene product . Faster bodyweight gain and fat accumulation in males may be associated to rapid thymic involution seen in male mice from 1 to 3 months of age . Both excessive caloric intake and obesity are proven to be factors that accelerate thymic involution [51, 52]. A 3-day in vitro culture of thymocytes revealed that DN1 cells from male mice exhibited a shorter half-life in comparison to females, suggesting that a possible difference in turnover time exists in primitive thymocytes in vivo .
Age-related thymic involution directly correlates with the declination of thymopoiesis as shown by sjTREC measurements in mice, human beings, and other primates [53–56]. In humans, females have significantly higher sjTREC levels than age-matched males aged 20–60 years old, with a maintained but less significant difference after the age of 60 . Based on our recent assay for sjTRECs in C57/BL6 mice, there is a similar trend between the different sexes. Data indicate that thymopoiesis level reflected by sjTREC value is quite variable between each individual mouse. The standard deviation of sjTREC values is larger in 1- and 3-month old mice than that in 7-month old mice. This agrees with the reports on humans that similarly show a wide variation of sjTREC values, particularly in younger cohorts . Despite variation of sjTREC levels in different mice, female mice contained a higher frequency of greater sjTRECs/μg value per microgram of thymocyte DNA at age 1 and 3 months in comparison to age-matched males (Fig. 2). This difference was not apparent at 7 months of age in mice, mirroring what has been observed in humans 60+ years of age . A greater rate of thymopoiesis may possibly explain why females have a higher absolute number of CD4 T-cell counts in comparison to men .
The recovery of thymopoiesis from myeloablative stem cell transplantation or intensive chemotherapy for curing cancer also proved to have an inverse relationship to age [58–60]. However a notable difference between males and females has not yet been recorded. Considering the key role of the thymus during specific therapies and the fact that there is a different involution rate between the female and male thymus, there would most likely be a differential thymopoietic recovery rate observed between the sexes. The likelihood is supported by the fact that castration alone or keratinocyte growth factor (KGF) plus an androgen blockage could enhance thymopoiesis after bone marrow transplantation [47, 61].
Despite all these findings, whether differences in thymic involution and thymopoiesis between males and females are solely a result of genetic variations, hormones, or both still remains a mystery. Whichever the case may be, future studies focusing on thymic involution and thymopoiesis regeneration should fully take sexual dimorphism into consideration. Failing to do so can readily lead to misinterpretation of results or futile therapeutic approaches that may be successful in one sex but not in the other. In addition to sex differences, one might also consider the variations in involution and thymopoiesis that may occur during specific windows of age, particularly from puberty to middle age, while in the process of determining prospective scientific studies that will be focused on this area of research.
Thymocyte development in the thymus requires a continual recruitment of non-self-renewable T-cell progenitors from bone marrow (BM) in addition to a functional thymic stroma that supports the differentiation and proliferation of thymocytes. There is a series of steps involved in the production of naive T-cell that are critical determinants of the immune system’s overall ability. These include steps at the levels of hematopoietic stem cells (HSCs), T-cell progenitors, the thymic niche, and the thymic stroma. Aging is accompanied by changes in these various steps .
Despite a great deal of debate, there is evidence to suggest that decreased lymphoid differentiation potential in HSCs occurs with aging . Mice receiving aged BM cells through transplantation have a lesser extent of thymic cellularity early after transplantation than those receiving BM cells from younger mice . However, these differences diminished 12 weeks after transplantation suggesting that intrinsic changes of HSCs in BM from aged individuals may not solely or contribute at all to age-related thymic atrophy. In addition, transfer of BM cells from young into aged mice did not lead to the restoration of thymic cellularity and architecture of the aged thymus .
Progenitors of T-cell are derived from HSCs originating in BM. The dosage of T-cell progenitors plays a role in determining thymic size and its capacity for thymopoiesis. The infusion of expanded T-cell progenitors co-cultured with OP9-DL1 cells enhances thymus engraftment and T-cell reconstitution . In addition, the MCL1 transgene induces an enlarged thymus in female mice by amplifying the pool of T-cell progenitors through enhancing their viability . These data imply that the decrease in T-cell progenitors in aged thymus could be the initiator of age-related thymic involution. Indeed, an experiment conducted in mice focusing on ckithi DN1 cells, which are early T-cell progenitors (ETPs), revealed that the ETP frequency and their potential to repopulate a fetal thymic organ culture decline in aged mice. Additionally, ETPs from aged mice exhibited less proliferation and more apoptosis . Other studies in mice also confirmed that a decrease in ETPs is a concomitant effect of age-related thymic involution [25, 67]. Even though there is a definite decrease in ETPs in aged mice, functional analyses have been inconsistent. Aspinall and Andrew demonstrated that DN1 cells from aged mice could repopulate an in vitro cultured fetal thymic organ normally . An in vivo experiment in which the fetal thymus was grafted under the renal capsule of both young and aged mice, showed no difference in thymic cellularity and absolute ETP number in the reconstituted fetal thymus 4–5 weeks after transplantation . These findings argue an intrinsic defect of ETP but suggest that the attrition and deterioration of ETPs in aged mice could be a secondary effect stemming from the insufficient extrinsic support due to dramatic loss of thymic stromal cells. This idea is reinforced by the observation that an intrathymic injection of ETPs sorted from young mice could not repopulate the atrophic thymus of an old recipient in the same way as they could reconstitute the thymus of a young recipient, which has an intact stroma .
Other than T-cell progenitors, the availability of thymic niche also plays a role in determining thymic size and thymopoiesis capability. An examination of thymic reconstitution after BM transplantation in various types of immunodeficient mice revealed an inverse relationship between thymus cell number and primitive DN precursor cell numbers . This elegant experiment demonstrated that even with the same overall thymic size and cellularity between IL-7 Rα−/− and RAG2−/− mice, transplantation of wild-type progenitor cells results in very different outcomes between these two strains. While RAG2−/− thymus was resistant to reconstitution even with a higher dose of BM cells, the IL-7 Rα−/− mice were restored to normal thymic cellularity. This can be attributed to the fact that the RAG2−/− thymus solely consists of DN cells whereas the IL-7 Rα−/− thymus contains much less DN cells while maintaining normal proportions of CD4+CD8+ double positive, CD4+ or CD8+ single positive, and DN cells. A greater number of primitive DN cells occupied the niche in the RAG2−/− thymus and formed a feedback that affected recruitment of wild-type progenitors. This is shown by pharmacological drugs, such as Sunitinib, which have the ability to enhance donor-derived cell engraftment in the thymus through the depletion of recipient-type ETPs for better niche accessibility . It is possible that changes in thymic architecture of aged subjects could have a negative influence on the niche availability, thus affecting recruitment of T-cell progenitors. However, this was not shown to be true experimentally. Injection of lineage-depleted BM cells of young mice into sub-lethally irradiated mice showed that the number of donor-derived DN1 cells did not differ between young and aged thymus 3–7 days after transplantation. In addition, the expression of progenitor homing factors, such as P-selectin and CCL25, did not differ between the young and aged thymic stromal tissues. However, 2–3 weeks following BM transplantation, the number of donor cells in a young thymus was more than 47-fold of those present in an old thymus. This is due to the fact that recruited progenitors undergo active proliferation and differentiation in the young thymus but stop further development in the old thymus . These results suggest that even though thymic stromal structure is diminished and disorganized with increased age, it is not a limiting factor for T-cell progenitor recruitment but is for further T-cell development that relies on optimal extrinsic support. It appears that the thymic niche accessibility could only play a role when strong thymopoiesis is initiated with sufficient extrinsic support from thymic stroma. This allows for a further developmental flow of recruited progenitors as, for example, when a young individual receives cytoreductive pre-conditioning for stem cell transplantation. In aging, however, instead of having a shortage of progenitor recruitment, the predominant defect stems from an insufficient support for progenitor survival, proliferation and differentiation attributed to an atrophic thymic stroma.
The thymic stroma has been analyzed extensively to show its significance in the development and proliferation of thymocytes. Stromal cells in the thymus are responsible for the production of various pro-thymic cytokines that are needed for sufficient extrinsic support for thymocyte development [72, 73]. The thymic stroma in aged subjects, particularly the TEC, deteriorates and exhibits a higher rate of apoptosis and lower rate of proliferation . A transgenic mouse with an N-terminal domain deletion in FOXN1, which regulates differentiation and proliferation of the TEC, stops the TEC differentiation at an intermediate progenitor stage. In this genetically modified mouse model, the thymus exhibited severe hypocellularity, lacking mature cortical or medullary regions. Although lymphocyte progenitor cells have the ability to enter the thymus, this population of cells appears to have defects in development . Further studies revealed that the FOXN1 gene is required for proper thymus formation and maintenance in both prenatal and postnatal stages, respectively, since a deletion of this gene resulted in thymic atrophy and significant medullary TEC loss at both stages [75, 76]. On the contrary, an overexpression of FOXN1 in thymic stroma led to an expansion of thymic epithelial cell numbers with higher proliferative rates in both young and aged mice, preventing an age-associated declination in thymopoiesis and changes in cortical and medullary thymic architecture . A recent study that used a conditional spontaneous gene deletion mouse model in which the expression of FOXN1 progressively declined with age, illustrated that there is a great loss of the TEC overtime. This loss is associated with other thymic aging phenotypes, all of which are notable at only 3–6 months of age. These aging phenotypes were partially rescued with an intrathymic injection of a FOXN1 plasmid . FOXN1 expression was found to progressively decrease in the thymic stroma tissue from normal aging mice , indicating that its gene function is closely associated with age-related thymic involution. The WNT signaling pathway, particularly WNT4, regulates FOXN1 expression and is predominately expressed in TECs . The expression of WNT4 also influences the quantity and organization of the TEC as early as thymic organogenesis. Deletion of the WNT4 gene led to fewer TECs per thymus in E15.5 day embryos with a similar difference observed in E18.5 day embryos. Frozen section-staining showed that the major loss of the TECs occurred in the K5+ medullary region. A graft of the Wnt4−/− fetal thymus beneath the renal capsule resulted in a much smaller thymus in comparison to the Wnt4+/+ fetal thymus graft, indicating that WNT4 is required for the development and maintenance of thymocytes both pre- and post-natally . Pharmacological studies show that WNT4 mRNA levels in TECs decrease with treatment of dexamethasone, suggesting that WNT4 plays a role in the protection of TECs against drug-induced thymic atrophy . Notably, a down-regulation of the expression of WNT4 in TECs was similarly observed in physiological aging mice . It is worthy to note that the growth of the thymic stroma is also regulated by the feedback stimulation of developing thymocytes. In SCID and RAG−/− mice, a block of thymocyte development at the DN3 stage results in improper thymic medullary structure. This feedback stimulation may be active at times when the thymus maintains better plasticity, as when mice are young or in embryonic developmental stages. As has been shown, the stromal structure of a RAG−/− fetal thymus grafted under renal capsule was completely restored by wild-type progenitors provided by recipients regardless of their ages . However, further evaluation is needed to determine how this interaction would occur in aged subjects that contain minimized thymic plasticity due to the deterioration of stromal structure. The aging induced atrophic thymic microenvironment itself would hinder thymocyte development, eliminating the possibility of feedback stimulation.
With a greater understanding of age-related thymic atrophy, several approaches for the restoration of thymopoiesis in an aged animal model were shown to be successful. The infusion of Ghrelin into 14-month old mice significantly increases thymopoiesis by increasing ETPs, Lin−sca-1+ckit+ BM cells, and ameliorating thymic stromal architecture . One can also restore a diminished ETP count in male mice through castration. Castration itself induces TEC growth and restores thymic stromal architecture [67, 84, 85]. Additionally, KGF promotes enhanced thymopoiesis in aged mice by inducing an increase of TECs, intrathymic interleukin-7 (IL-7) production, and a reorganization of cortical and medullary architecture . In a young mouse model, a report showed that enhanced thymopoiesis is accompanied by an increased entry of ETPs after androgen withdrawal through CCL25 which is induced by TEC proliferation . In all of the above cases, increased thymopoiesis requires a pre-existing ground of enhanced thymic stroma which includes not only a greater number of TECs but also the restoration of the cortical and medullary architecture. This might explain why local increases in intrathymic IL-7 levels could only promote the transition from DN1 to DN2 in aged mice, but not increase T-cell output since there was a lack of an enhanced thymic stroma . This is reminiscent of what was found in MCL1 transgenic mice. By promoting the viability of ETPs, the MCL1 transgene induced an overall enlarged thymus with proportional expanded TECs which possibly is a result of an early embryonic interaction with progenitors. However, mouse BM transplantation experiments show that fortified progenitors with MCL1 transgene do not have the ability to produce an enlarged thymus in wild-type recipients . This may suggest that a capacity-matched thymic stroma is first required for the development of enhanced progenitors. In other words, the capability of thymic stroma keeps check of thymic size. These results highlight the importance of the thymic stroma in guiding thymocyte development. Collectively, HSCs, ETPs, thymic niche, and thymic stroma all contribute to the overall capability of the thymus and must be considered at different steps in the restoration of age-related thymic involution.
Thymus size and function are determined not only by genetics but also by early life nutrition and other stochastic events. Both pre-natal and early post-natal periods are critical windows for thymus development. Influences during these periods can lead to long-term consequences into adulthood including altered cell-mediated immunity or accelerated thymic aging. Sex differences in age-related thymic involution and the capacity of thymopoiesis are particularly obvious when there is a strong activation of sex hormones as there is from puberty to middle age. The efficacy of possible strategies that restore thymopoiesis should be assessed in both sexes due to dimorphic differences. Age-related thymic involution results in the progressive deterioration of the thymic stromal microenvironment particularly due to a loss of TECs. Measurements that aim to improve thymopoiesis and restore T-cell function in the elderly should first explore the possibility to expand thymic stroma and ameliorate the thymic microenvironment. Several genetically modified mouse models may suggest the roles that particular genes play in thymic aging. It is worthy to note that age-related thymic atrophy is accompanied by a systematic change in expression profiles of a variety of genes . This is true not only in thymic stromal cells  but also in thymocytes . Recent experiments comparing global thymic stromal gene profiles of young, old, and old castrated mice revealed that castration induced a transient overall restoration of thymic cellularity without changing gene expression profiles which were maintained the same as that of an aged thymic stroma . This implies that thymic aging is a persistent degenerative change that might not be completely restored qualitatively. A study showed that deleting tumor suppressor expression induced cancer but attenuated thymic involution . Thus, it is possible that age-related thymic aging has its own evolutionary benefit by minimizing the incidence of tumors through counteracting other aging associated alterations like mutations .