Водородът засилва имунитета при рак на бял дробНаучно Изследване
оригинално заглавие (букв. прев.): Две седмици вдишване на водород могат значително да обърнат адаптивното и вродено стареене на имунната система на пациенти с напреднал недребноклетъчен рак на белия дроб: самоконтролирано проучване
1 Fuda Cancer Hospital of Jinan University; Fuda Cancer Institute, Guangzhou, Guangdong Province, China
2 Fuda Cancer Hospital of Jinan University, Guangzhou, Guangdong Province, China
3 Central Laboratory, Peking University Cancer Hospital, Beijing, China
Публикувано на: 25/12/2020
След стандартни лечения, традиционният модел за повишаване на противотуморния имунитет включва извършване на имунна реконституция (напр. приемни имунни клетъчни терапии или лекарства за повишаване на имунитета) за предотвратяване на рецидив.
За пациенти с напреднал недребноклетъчен рак на белия дроб, ние докладваме тук за две цели, имуносресценцията за напреднал недребноклетъчен рак на белия дроб и вдишване на водороден газ за възстановяване на имунитета.
От 1 юли до 25 септември, 2019 г., 20 пациенти с недребноклетъчен рак на белия дроб бяха включени за оценка на имуноснесценцията на подгрупи от лимфоцити в периферната кръв, включително Т клетки, естествени убийци/естествени убийци Т клетки и гама делта Т клетки. По време на периода на изчакване за преглед, свързан с лечението, беше извършена две седмици инхалация на водород.
Всички пациенти са вдишвали смес от водород (66,7%) и кислород (33,3%) със скорост на газовия поток от 3 L/min в продължение на 4 часа всеки ден.
Нито един от пациентите не е получавал стандартно лечение по време на периода на вдишване на водород.
След тестване преди лечение се наблюдават основни индекси на имуноснесценция. Ненормално по-високите индекси включват изчерпани цитотоксични Т клетки, старещи цитотоксични Т клетки и убийци Vδ1 клетки. След 2 седмици водородна терапия, броят на изчерпаните и стареещи цитотоксични Т клетки намалява до нормалните граници и има увеличение на Vδ1 клетките убийци.
Ненормално по-ниските индекси включват функционални помощни и цитотоксични Т клетки, Th1, общи естествени Т-клетки убийци, естествени убийци и Vδ2 клетки.
След 2 седмици терапия с водород, всичките шест клетъчни подгрупи се увеличават до нормалните граници.
Настоящите данни показват, че имуноснесценцията на напреднал недребноклетъчен рак на белия дроб включва почти всички лимфоцитни подгрупи и 2 седмици лечение с водород може значително да подобрят повечето от тези индекси.
Проучването е одобрено от Комитета по етика на болницата за рак Фуда, университета Джинан в Китай (одобрение № Fuda20181207) на 7 декември2018 г. и е регистриран на ClinicalTrials.gov (ID: NCT03818347) на 24 януари 2019 г.
The recurrence and metastasis of malignant tumors are difficult to control, and the main reasons can be attributed to three: 1) immune aging; 2) tumor-related factors; 3) cancer treatment itself.  Immunosenescence is characterized by the gradual degeneration of the immune system during natural aging. Prolonged tumor-bearing status also leads to depletion and senescence of cytotoxic lymphocytes, including T cells, natural killer (NK) cells, NKT cells, and γδ (γδ) T cells. ,  are tumor factors associated with excessive tumor burden and severe suppression of immune function, including increased CD4+CD25+ regulatory T cells.  Currently, various conventional treatments have been shown to accelerate the recurrence and metastasis of residual tumors, including surgery,  radiotherapy,  chemotherapy,  and even general anesthesia. 
Lung cancer is associated with the highest global incidence of all cancers, with non-small cell lung cancer (NSCLC) accounting for approximately 85% of cases.  Due to the lack of early symptoms of lung cancer, many patients are diagnosed at an advanced stage, and survival is usually 1 year.  Studies have shown that nearly 27% of patients undergoing surgery and chemotherapy die from recurrence and metastasis.  Therefore, before and after standard therapy, immune reconstitution is a necessary approach to prolong survival in patients with advanced cancer. .Adoptive immune cell replenishment, particular or general immunological activators, cancer vaccinations, and exercise are the main techniques for immune rebuilding now being used.  Hydrogen can permeate into lymphocyte mitochondria as an antioxidant gas and specifically scavenge oxygen free radicals. , Additionally, it has been demonstrated that hydrogen inhalation decreases the expression of PD-1, a sign of cell exhaustion, on the surface of cytotoxic T lymphocytes in patients with advanced colorectal cancer, extending both progression-free survival and overall survival.  In this study, immunosenescence in advanced NSCLC patients was examined, and immunosenescence was reversed while the patients were waiting for therapy using hydrogen inhalation.
Subjects and Methods
This self-controlled study included 20 advanced NSCLC patients admitted at Fuda Cancer Hospital between July and September 2019 who had to wait two weeks for therapy. The following inclusion criteria were listed in the clinical trial that was registered (ClinicalTrials.gov, ID: NCT03818347; Registration Date: January 24, 2019): stage III or IV NSCLC diagnosed by imaging and pathology; tumor number 1-6; maximum tumor length 2 cm; Karnofsky performance status (KPS) score 7019; expected survival time > 6 months; platelet count 80 109/L; white blood cell count 3 Patients having a cardiac pacemaker, brain metastases, grade 3 hypertension, diabetic complications, and severe cardiac and pulmonary dysfunction were excluded from the study. During the duration of hydrogen inhalation, none of the patients received any conventional care.
In order to guarantee that any concerns regarding the accuracy or integrity of any component of the work are duly investigated and addressed, the writers are responsible for all aspects of the work. On December 7, 2018, the Fuda Cancer Hospital’s Ethics Committee granted their permission for the study protocol (approval number Fuda20181207). Each participant gave their written, informed consent in accordance with the Declaration of Helsinki.
Hydrogen inhalation method
Hydrogen was delivered by a hydrogen-oxygen nebulizer (AMS-H-03, Shanghai Asclepius Meditec Co., Ltd., Shanghai, China). The patients remained situated or prostrate, and breathed in a blend of hydrogen (66.7%) and oxygen (33.3%) by means of a nasal tube or veil with a gas stream rate of 3 L/min. The hydrogen inward breath was proceeded for 4 hours per day for 2 weeks in a specialized treatment facility.
Pulmonary symptoms and KPS score
Before and after hydrogen therapy, the respiratory function was assessed by very experienced respiratory physicians using a pulmonary function tester (Autospiro AS-507; Minato Medical Science, Tokyo, Japan), pulmonary tumor-related symptoms and the KPS score were evaluated by resident doctors in charge.
Roughly 5 mL of fringe blood was extricated from elbow of all selected patients some time recently hydrogen inward breath, as well as at one and two weeks after hydrogen inward breath. Fringe blood monocyte cells were separated employing a Ficoll arrangement and labeled with fluorescent antibodies (BD Biosciences, San Jose, CA, USA). T lymphocytes, NK, NKT, and γδ T cells were analyzed utilizing stream cytometry (FACSanto II; BD Biosciences, San Jose, CA, USA) by a proficient third-party assessment center (Shuangzhi Purui Therapeutic Research facility Co., Ltd., Wuhan, Hubei Area, China).
The calculation strategy of test estimate is based on MedSci Test Measure instruments (MSST) computer program (MedSci, Shanghai, China). Pneumonic indications some time recently and after hydrogen inward breath were compared utilizing combined t-test. Respiratory work, KPS score, and immunoassays were compared by rehashed measures investigation of change and Bonferroni’s different comparison test. Measurable contrasts were demonstrated by P < 0.05. All examinations and figures were delivered utilizing GraphPad Crystal 5.0 (GraphPad computer program, San Diego, CA, USA).
Clinical data of NSCLC patients
A total of 20 consecutive patients admitted to the hospital participated in this study. The clinicopathological data are listed in [Table 1].
Changes in the KPS score and pulmonary symptoms of NSCLC patients
All patients received hydrogen inhalation therapy for 2 weeks and experienced no treatment-related symptoms or side effects. The results of respiratory function, KPS score, and pulmonary symptoms at 2 weeks before and after hydrogen therapy are shown in the table [Table 1]. 2] as shown in the figure. At the end of hydrogen treatment, there were fewer people with different pulmonary symptoms and a statistically lower number of moderate coughs and mild shortness of breath.
Changes in T cell subsets of NSCLC patients undergoing hydrogen therapy
This part of the test consists of three components: CD4+ T cells, CD8+ T cells, and cytokine-secreting helper T (Th) cells. Six major markers of immunosenescence were identified prior to starting hydrogen therapy. Among these indices, abnormally low indices included percentages of functional helper and cytotoxic T cells, Th1, and follicular Th cells; abnormally high indices were associated with percentages of depleted and aged cytotoxic T cells [Figure 1] ]. In the CD4+ subgroup, functional Th cells returned to the normal range after 1 week of hydrogen inhalation, and increased significantly after 2 weeks (P<0.05).After 2 weeks of hydrogen therapy, both depleted Th cells and regulatory T cells were gradually decreased (both P<0.05) [Fig. 1]A. In the CD8+ subgroup, functional cytotoxic T cells returned to the normal range after 2 weeks of hydrogen inhalation (P<0.05); after 2 weeks of treatment, both exhausted and senescent cytotoxic T cells gradually returned to the normal range (both P<0.05). 0.05) [Figure 1]B. Among the cytokine-secreting Th subsets, only Th1 cells gradually increased close to the normal range after 2 weeks of hydrogen inhalation treatment (P < 0.05; [Fig. 1] C. There were no significant changes in other cells within 2 weeks after hydrogen inhalation treatment.
Figure 1: Immunoassays of T cell subsets before and after hydrogen inhalation in patients with non-small cell lung cancer. NOTE: (A) Test results for CD4+ subsets. (B) Test results of CD8+ subsets. (C) Test results of cytokine-secreting CD4+ subsets. The red parallel long line in the figure represents the normal range, the black short line represents the average value at each time point, and the pink cell name represents the abnormal index before hydrogen treatment. Bonferroni’s multiple comparison test analysis. *P<0.05. CXCR5: C-X-C chemokine receptor type 5; Th: helper T; Tfh: follicular helper T.
Changes in NKT and NK cells of NSCLC patients undergoing hydrogen therapy
The percentages of total NKT cells, activated NK, and killer NK subsets were below the normal range before starting hydrogen therapy [Figure 2]. After one week of hydrogen inhalation, the activated NK cell subset increased to the normal range [Figure 2]C. After two weeks, both total NKT and killer NK subsets were higher than pretreatment percentages (both P<0.05; [Fig. 2] A and [Fig. 2] D, and activated NK cells were significantly increased (P < 0.01; [Fig. 2]). ] ] C. The total number of NK cells did not change significantly within 2 weeks after hydrogen inhalation [Fig. 2] B.
Figure 2: Immunoassay of natural killer (NK)T and NK cells before and after hydrogen inhalation in non-small cell lung cancer patients.
Note: (A) Changes in the number of NKT cells. (B–D) Test results of the NK subsets. The parallel red long lines in the figure represent the normal range, the black short lines represent the average value at each time point, and the pink cell names represent the abnormal indicators before hydrogen treatment. Data were analyzed by repeated measures analysis of variance followed by Bonferroni’s multiple comparison test. *P < 0.05, **P < 0.01.
Changes in γδ T cell subsets of NSCLC patients undergoing hydrogen therapy
Before starting hydrogen therapy, the percentages of Vδ1 cells and killer Vδ1 cells were above the normal range, and the percentage of Vδ2 cells was below the normal range [Figure 3]. After 1 week of hydrogen inhalation, the percentage of Vδ2 cells in γδT cells increased to the normal range [Fig. 3]C. After two weeks, the percentages of total γδT cells and Vδ2 cells were higher than those before treatment (both P<0.05; [Fig. 3]A and [Fig. 3]C, depleted Vδ2 cells were lower than those before treatment (P < 0.05; [Fig. 3]C; Depleted Vδ1 cells significantly decreased (P<0.001; [Fig. 3]B and Killer Vδ2 cells were significantly increased (P<0.01; [Fig. 3]C) other cells were not significantly increased under hydrogen for 2 weeks). change) inhalation.
Figure 3: Immunoassay of the gamma delta (γδ) T cell subsets before and after hydrogen inhalation in non-small cell lung cancer patients.
Note: (A) Change in the number of γδ T cells. (B) Test results of the Vδ1 subsets. (C) Test results of the Vδ2 subsets. The parallel red long lines in the figure represent the normal range, the black short lines represent the average value at each time point, and the pink cell names represent the abnormal indicators before hydrogen treatment. Data were analyzed by repeated measures analysis of variance followed by Bonferroni’s multiple comparison test. *P < 0.05, **P < 0.01, ***P < 0.001. NK: Natural killer ; PD-1: programmed cell death protein 1.