## 4 Discussion  
### Main Discussion  
Erythrocyte aggregation is an intrinsic property of native blood that has a direct effect on blood viscosity and circulation, especially at low shear stress. The aggregation of erythrocytes is determined by intrinsic (e.g., cell surface charge) and extrinsic (e.g., plasma fibrinogen concentration, hematocrit, shear rate) factors [26]. This study first demonstrated that an acute 12% oxygen hypoxia exercise test augmented erythrocyte aggregation, which was mainly mediated by the intrinsic factors of erythrocytes. Furthermore, HE had a superior effect to NE in ameliorating cardiopulmonary capacity. After 6 weeks of interventions, only the HE further promoted aggregation via both intrinsic aggregability and fibrinogen-erythrocyte interaction, which was accomplished with accumulating counts of stress reticulocytes and erythrocyte senescence. We further demonstrated that NE combined with either decreased fibrinogen binding factor or increased intrinsic aggregability, which seems to be the rationale as to why NE did not alter the aggregation reaction after 6 weeks. Hemoconcentration is one of the most considerable factors in increasing blood aggregation [27], which increases the opportunity for cell-to-cell contact and largely explains the elevated aggregation during exercise [28]. However, aggregation was not different for suspensions with standardized hematocrit, indicating that hemoconcentration was less relevant to the increased aggregation caused by an acute HET. In contrast to the acute effect,  
Figure 5: Measurement of erythrocyte composition: (**A**) the percentage of reticulocytes, (**B**) the percentage of stress reticulocytes, and (**C**) the percentage of mature erythrocytes. NE, normoxia training group; HE, hypoxia training group; CTL, control group; Pre, pretraining: Post, post-training; Rt, at rest; HET, immediately after HET; * \(p\) < 0.05, Rt vs. HET; \(\tau\)\(p\) < 0.05, pre vs. post. Values are the mean \(\pm\) SEM. hemodilution, a usual long-term effect of regular exercise, was not found in either HE or NE; erythrocyte and hemoglobin counts and hematocrit levels were not apparently altered with either exercise regimen. Hypoxia and physical exercise are independent of each other and are both highly potent metabolic stressors. Combined, these factors significantly elevated the degree of whole blood aggregation, as we noticed after acute HET and HE. Hypoxia can stimulate EPO secretion under the regulation of HIF-1 [15]. However, a severe hypoxic condition or acute exercise may enhance proinflammatory factor levels, such as those of MPO, TNF-\(\alpha\), IL-1, or IL-6. This status may inhibit the EPO response and hematological adaptations [29]. With inflammation, an elevated fibrinogen concentration is mainly responsible for enhancing aggregation [7]. In the clinical setting, a patient with exertional periodic breathing resembles the model of intermittent hypoxia exposure, likely triggering increased ROS [30] and proinflammatory status [31], thus further deteriorating erythrocyte aggregation [32]. Interestingly, neither normoxic exercise training alone nor long-term hypoxic exposure alone was sufficient to suppress those proinflammatory cytokine effects [33]. However, the 15% oxygen exercise training used in the present study has been demonstrated to suppress MPO and IL-6 production and lipid peroxidation caused by severe exercise or acute hypoxia [34]. Although the plasma fibrinogen concentration responded to the 15% oxygen exercise training, further investigation is still needed. In contrast to the NE, the HE dramatically increased ROS during hypoxic training, presumed to further lead to CD47 and CD147 downregulation and enhanced erythrocyte aggregation [35]. Elevated ROS levels are speculated to stabilize HIF-1 production [36]. We previously pointed out that HE can strongly deteriorate erythrocyte antioxidative capacity [13]. Upon return to normoxia, there was an excessive accumulation of ROS in reticulocyte mitochondrial mass [37].