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A single heterochronic blood exchange reveals rapid inhibition of multiple tissues by old blood

View Article: PubMed Central - PubMed

ABSTRACT

Heterochronic parabiosis rejuvenates the performance of old tissue stem cells at some expense to the young, but whether this is through shared circulation or shared organs is unclear. Here we show that heterochronic blood exchange between young and old mice without sharing other organs, affects tissues within a few days, and leads to different outcomes than heterochronic parabiosis. Investigating muscle, liver and brain hippocampus, in the presence or absence of muscle injury, we find that, in many cases, the inhibitory effects of old blood are more pronounced than the benefits of young, and that peripheral tissue injury compounds the negative effects. We also explore mechanistic explanations, including the role of B2M and TGF-beta. We conclude that, compared with heterochronic parabiosis, heterochronic blood exchange in small animals is less invasive and enables better-controlled studies with more immediate translation to therapies for humans.

No MeSH data available.


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Levels of B2M in young muscle and brain correlate positively with the heterochronicity of blood exchange.(a) Muscle cryosections of 10 μm and 25 μm brain-SGZ cryosections of isochronically and heterochronically apheresed mice (that had experimental muscle injury) were immuno-stained for B2M and counter-stained for Hoechst to label all nuclei. Representative images were acquired at the sites of muscle injury (Mu in) outside the injury-repair (Mu out) and at the hippocampi-DG areas (brain DG), scale bar is 50 μm for muscle and liver, and 100 μm for brain. (b) Pixel density of B2M was quantified using Image J from serial cryosections represented in a; and shown are the means and standard errors. In muscle: ***,**P<0.005. Significant differences were observed between YY and YO (P=0.004), OY and OO (0.001), YO and OY (P=0.0007), and YY and OO (P=0.006), N=5–7 per group. In brain: ****P<0.00005. Significant differences were observed between YY and YO (P=0.00001), and YY and OO (P=0.004). (c) Western SDS–polyacrylamide gel electrophoresis was used to analyse B2M levels in one microlitre of cell-free blood serum from 5 young (Y) and 5 old (O) mice. ECL images were quantified by ImageJ and expressed as background-corrected pixel volume. N=5. P=0.5. B2M becomes increased with age in muscle and brain but it is not elevated in old blood serum as compared with young. After heterochronic blood exchange B2M is increased by old blood in young muscle and decreased by young blood in old muscle (regionally, outside of the injury site). B2M is also increased in young hippocampi-DG after exchange with old blood, but B2M is not diminished in the old DG after young blood exchange. Shown are means±s.e.m. for all histograms.
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f4: Levels of B2M in young muscle and brain correlate positively with the heterochronicity of blood exchange.(a) Muscle cryosections of 10 μm and 25 μm brain-SGZ cryosections of isochronically and heterochronically apheresed mice (that had experimental muscle injury) were immuno-stained for B2M and counter-stained for Hoechst to label all nuclei. Representative images were acquired at the sites of muscle injury (Mu in) outside the injury-repair (Mu out) and at the hippocampi-DG areas (brain DG), scale bar is 50 μm for muscle and liver, and 100 μm for brain. (b) Pixel density of B2M was quantified using Image J from serial cryosections represented in a; and shown are the means and standard errors. In muscle: ***,**P<0.005. Significant differences were observed between YY and YO (P=0.004), OY and OO (0.001), YO and OY (P=0.0007), and YY and OO (P=0.006), N=5–7 per group. In brain: ****P<0.00005. Significant differences were observed between YY and YO (P=0.00001), and YY and OO (P=0.004). (c) Western SDS–polyacrylamide gel electrophoresis was used to analyse B2M levels in one microlitre of cell-free blood serum from 5 young (Y) and 5 old (O) mice. ECL images were quantified by ImageJ and expressed as background-corrected pixel volume. N=5. P=0.5. B2M becomes increased with age in muscle and brain but it is not elevated in old blood serum as compared with young. After heterochronic blood exchange B2M is increased by old blood in young muscle and decreased by young blood in old muscle (regionally, outside of the injury site). B2M is also increased in young hippocampi-DG after exchange with old blood, but B2M is not diminished in the old DG after young blood exchange. Shown are means±s.e.m. for all histograms.

Mentions: To start looking into the molecular mechanisms that are responsible for these rapid influences of circulation on tissue repair and maintenance, we studied the levels of B2M. B2M is the invariant chain of MHC class I that becomes elevated with inflammation and based on current reports is over-pronounced in old muscle and brain, as compared with young81420. B2M levels were assayed by immunofluorescence in tissue cryosections (Fig. 4a,b) and by western blotting (Supplementary Fig. 5) in the young and old mice that underwent isochronic versus heterochronic blood exchange as described above. For tissues derived from mice injured with CTX in their TAs, the immunofluorescence on muscle and brain tissue cryosections demonstrated that exchange with old blood rapidly (within 6 days), elevated the B2M levels in young muscle located outside of the CTX injury, and in the SGZ of the young hippocampus, (Fig. 4a,b). Interestingly, the B2M remained high in the old hippocampi of the heterochronically exchanged animals (Fig. 4a,b). Furthermore, for muscle these age-specific differences in B2M were less pronounced between YY and YO cohorts and were undetectable between the OO and OY cohorts when immunofluorescence was performed at the sites of CTX injury—muscle regeneration (Supplementary Fig. 5) in agreement with the previous findings that inflammation overlaps in space and time with muscle repair and that some degree of transient inflammation is needed for successful myogenesis721.


A single heterochronic blood exchange reveals rapid inhibition of multiple tissues by old blood
Levels of B2M in young muscle and brain correlate positively with the heterochronicity of blood exchange.(a) Muscle cryosections of 10 μm and 25 μm brain-SGZ cryosections of isochronically and heterochronically apheresed mice (that had experimental muscle injury) were immuno-stained for B2M and counter-stained for Hoechst to label all nuclei. Representative images were acquired at the sites of muscle injury (Mu in) outside the injury-repair (Mu out) and at the hippocampi-DG areas (brain DG), scale bar is 50 μm for muscle and liver, and 100 μm for brain. (b) Pixel density of B2M was quantified using Image J from serial cryosections represented in a; and shown are the means and standard errors. In muscle: ***,**P<0.005. Significant differences were observed between YY and YO (P=0.004), OY and OO (0.001), YO and OY (P=0.0007), and YY and OO (P=0.006), N=5–7 per group. In brain: ****P<0.00005. Significant differences were observed between YY and YO (P=0.00001), and YY and OO (P=0.004). (c) Western SDS–polyacrylamide gel electrophoresis was used to analyse B2M levels in one microlitre of cell-free blood serum from 5 young (Y) and 5 old (O) mice. ECL images were quantified by ImageJ and expressed as background-corrected pixel volume. N=5. P=0.5. B2M becomes increased with age in muscle and brain but it is not elevated in old blood serum as compared with young. After heterochronic blood exchange B2M is increased by old blood in young muscle and decreased by young blood in old muscle (regionally, outside of the injury site). B2M is also increased in young hippocampi-DG after exchange with old blood, but B2M is not diminished in the old DG after young blood exchange. Shown are means±s.e.m. for all histograms.
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f4: Levels of B2M in young muscle and brain correlate positively with the heterochronicity of blood exchange.(a) Muscle cryosections of 10 μm and 25 μm brain-SGZ cryosections of isochronically and heterochronically apheresed mice (that had experimental muscle injury) were immuno-stained for B2M and counter-stained for Hoechst to label all nuclei. Representative images were acquired at the sites of muscle injury (Mu in) outside the injury-repair (Mu out) and at the hippocampi-DG areas (brain DG), scale bar is 50 μm for muscle and liver, and 100 μm for brain. (b) Pixel density of B2M was quantified using Image J from serial cryosections represented in a; and shown are the means and standard errors. In muscle: ***,**P<0.005. Significant differences were observed between YY and YO (P=0.004), OY and OO (0.001), YO and OY (P=0.0007), and YY and OO (P=0.006), N=5–7 per group. In brain: ****P<0.00005. Significant differences were observed between YY and YO (P=0.00001), and YY and OO (P=0.004). (c) Western SDS–polyacrylamide gel electrophoresis was used to analyse B2M levels in one microlitre of cell-free blood serum from 5 young (Y) and 5 old (O) mice. ECL images were quantified by ImageJ and expressed as background-corrected pixel volume. N=5. P=0.5. B2M becomes increased with age in muscle and brain but it is not elevated in old blood serum as compared with young. After heterochronic blood exchange B2M is increased by old blood in young muscle and decreased by young blood in old muscle (regionally, outside of the injury site). B2M is also increased in young hippocampi-DG after exchange with old blood, but B2M is not diminished in the old DG after young blood exchange. Shown are means±s.e.m. for all histograms.
Mentions: To start looking into the molecular mechanisms that are responsible for these rapid influences of circulation on tissue repair and maintenance, we studied the levels of B2M. B2M is the invariant chain of MHC class I that becomes elevated with inflammation and based on current reports is over-pronounced in old muscle and brain, as compared with young81420. B2M levels were assayed by immunofluorescence in tissue cryosections (Fig. 4a,b) and by western blotting (Supplementary Fig. 5) in the young and old mice that underwent isochronic versus heterochronic blood exchange as described above. For tissues derived from mice injured with CTX in their TAs, the immunofluorescence on muscle and brain tissue cryosections demonstrated that exchange with old blood rapidly (within 6 days), elevated the B2M levels in young muscle located outside of the CTX injury, and in the SGZ of the young hippocampus, (Fig. 4a,b). Interestingly, the B2M remained high in the old hippocampi of the heterochronically exchanged animals (Fig. 4a,b). Furthermore, for muscle these age-specific differences in B2M were less pronounced between YY and YO cohorts and were undetectable between the OO and OY cohorts when immunofluorescence was performed at the sites of CTX injury—muscle regeneration (Supplementary Fig. 5) in agreement with the previous findings that inflammation overlaps in space and time with muscle repair and that some degree of transient inflammation is needed for successful myogenesis721.

View Article: PubMed Central - PubMed

ABSTRACT

Heterochronic parabiosis rejuvenates the performance of old tissue stem cells at some expense to the young, but whether this is through shared circulation or shared organs is unclear. Here we show that heterochronic blood exchange between young and old mice without sharing other organs, affects tissues within a few days, and leads to different outcomes than heterochronic parabiosis. Investigating muscle, liver and brain hippocampus, in the presence or absence of muscle injury, we find that, in many cases, the inhibitory effects of old blood are more pronounced than the benefits of young, and that peripheral tissue injury compounds the negative effects. We also explore mechanistic explanations, including the role of B2M and TGF-beta. We conclude that, compared with heterochronic parabiosis, heterochronic blood exchange in small animals is less invasive and enables better-controlled studies with more immediate translation to therapies for humans.

No MeSH data available.


Related in: MedlinePlus