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Blood parameters before and after hydrogen treatment. (A) Red blood cell counts and hemoglobin levels; the red line in the represents the lower limit of the reference range. (B) Total protein and albumin concentrations; the red lines in the figures represent the lower limit of the reference range. (C) White blood cell and lymphocyte counts; the red lines in the figures represent the reference range.
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Moreover, these two indexes began to increase continuously one month later (Figure 1A), and the blood transfusion was completely stopped after two months. The level of serum total protein and albumin began to rise after supportive treatments and continued to increase after hydrogen inhalation, reaching a normal range after two and a half months, at which time, supplements were no longer required (Figure 1B). The leukocyte count decreased to the upper limit of the reference range after antibiotic treatment and continued to decrease after hydrogen inhalation, falling to normal levels after 1.5 months (Figure 1C left). The total number of lymphocytes was low at the time of admission, and the number began to rebound after one and a half months of hydrogen inhalation, returning to normal after two and a half months (Figure 1C right).
Changes in the tumor markers before and after hydrogen treatment. The red lines in the figures represent the upper limit of the reference ranges.
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Following hydrogen inhalation, the three markers did not decline and the AFP even exceeded the level at the time of admission; however, these levels rapidly declined afterwards, and all fell to within the normal reference range after two and a half months (Figure 2).
(A) CT imaging findings at different time-points before and after hydrogen treatment. In the first line of figures, the triangle and the contour lines represent the location and range of the gallbladder. In the second line of figures, the contour lines represent the location and range of the liver metastases. In the third line of figures, the arrows represent the location of the duodenal fistula or obstruction. (B) Evaluation of the effect of hydrogen treatment at different time-points according to CT imaging and RECIST 1.1.. Abbreviations: PD, progressed disease; SD, stable disease; PR, partial remission; CT, computed tomography.
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Abdominal CT images were analyzed at different time points before and after hydrogen treatment (Figure 3A). After hydrogen treatment for: 1) one month, the sum was 16.7 cm (37% increase compared with pretreatment, PD); 2) two and a half months, the sum was 12.9 cm (6% increase compared with pretreatment, SD); and 3) six months, the total was 8.1 cm (33% decrease compared with pretreatment, PR) (Figure 3B).
In terminally differentiated CD8+ T cells, the proportion of exhausted cells varied with the treatment time. The results were tested by flow cytometry, in which the terminally differentiated CD8+ T cells were labeled CD3+CD8+CD27-, in which PD-1+, was considered a marker of exhaustion and non-exhausted cells were PD-1.
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In the terminally differentiated CD8+ T cells, the proportion of exhausted cells was 46.8% at two weeks before treatment, 31.3% at six months after treatment, and 27.6% after nine months (Figure 4).