CN1300594A - Ginsengdiol histsaponin and application of its composition in treating hematopathy - Google Patents
Ginsengdiol histsaponin and application of its composition in treating hematopathy Download PDFInfo
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Abstract
The application of ginsengdiol histsaponin and its composition in treating hematopathy is disclosed. Said composition is composed of ginsengdiol histsaponin, medicinal carrier and/or medicinal exipitent, and can stimulate the reproduction of hematopoietic cells. Its advantages include high curative effect, no toxic by-effect and low cost.
Description
The present invention relates to the hematopathy clinical treatment, mainly is that Ginsengdiol histsaponin among the ginsenoside and compositions thereof are applied to hemopathic clinical treatment.
At present both at home and abroad to intractable hematopathy, still do not have specific treatment because of its cause of disease complexity, with prednisone and androgenic routine treatment well-known side effect is arranged for a long time, and the recombinant growth factors therapy costs an arm and a leg, can't conventionally use, and invalid to a lot of intractable hematopathys.Radix Ginseng is as defying age analeptic and existing over thousands of year history of qi and blood tonifying medicament, Radix Ginseng extract is a Radix Ginseng total saponins, to the existing many reports of nerve, cardiovascular, endocrine, immunity and antineoplastic experimentation, but seldom to the experimentation of blood aspect, especially Ginsengdiol histsaponin and compositions thereof are not seen the clinical report that this respect is arranged yet.
The purpose of this invention is to provide a kind of ginsenoside of containing, especially the compositions of Ginsengdiol histsaponin is applied to hemopathic clinical treatment.
Compositions provided by the invention is by the ginsenoside, and especially Ginsengdiol histsaponin and pharmaceutical carrier and/or pharmaceutical excipient are formed, and are applied to hemopathic treatment.
The present invention is with the ginsenoside, especially Ginsengdiol histsaponin and compositions thereof, be applied to hemopathic treatment, curative effect is better than the existing conventional medicine, and does not have toxicity in treatment, compositions provided by the invention, hematopoietic cell had the effect of stimulating proliferation, hemopoietic progenitor cell to Patients with Aplastic Anemia has the effect of stimulating proliferation, and the leukemia CFU-GM is had the two-way function of inhibition of proliferation and stimulation, the effect that has stronger promotion chemotherapeutic to kill and wound the leukaemia.
Ginsenoside provided by the invention, especially the compositions of Ginsengdiol histsaponin, it is big to have overcome the toxic and side effects that existing treatment hematopathy medicine exists, and weak point such as cost an arm and a leg, for treating hematopathy provides new approach, expanded ginsenoside's clinical application range.Alleviated the misery that the patient brings because of toxic and side effects simultaneously,, also alleviated hematopathy patient's financial burden especially because the said composition cost is lower.
The present invention is described further by following examples.
Embodiment 1: a kind of prescription of Ginsengdiol histsaponin compositions
Prescription: panoxadiol's powder 30 gram dextrin 470 are restrained into 1000
Preparation: it is an amount of to get fibrous root of Radix Ginseng coarse powder (20 mesh sieve), carries out percolation with debita spissitudo ethanol and is extracted into colourlessly, collects extracting solution, leave standstill more than 24 hours, get supernatant and reclaim ethanol, medicinal liquid is concentrated into debita spissitudo, upper prop to there not being the alcohol flavor, carry out separation and Extraction with the column chromatography for separation method, separate with low ethanol, collect separation and Extraction liquid, through vacuum drying after a few hours, get crystalloid, faint yellow panoxadiol's elaboration.Take by weighing elaboration 30 grams, grind into impalpable powder, cross 80 mesh sieves,, incapsulate and get final product every capsules 0.5 gram (containing 30 milligrams of the pure product of panoxadiol) with the dextrin mix homogeneously.Quality standard meets Ministry of Public Health medicine sanitary standard regulation and Chinese Pharmacopoeia 95 editions.
Embodiment 2: Ginsengdiol histsaponin is studied the effect of mouse bone marrow cells hemopoietic progenitor cell
Radix Ginseng is as medicinal existing several thousand history, the Radix Ginseng total saponins that its effective ingredient mainly is made up of more than 20 kinds of saponin monomers, and the pharmacological action of each composition is not quite similar.The ginsenoside is divided into Ginsengdiol histsaponin (Pananxadiol Saponin by the contained amount of hydroxyl groups difference of saponin monomer unit; PDS), the panaxatriol organizes Saponin (Panaxtrol Saponin; PTS) two big classes.Hemopoietic progenitor cell In vitro culture technology is adopted in this research, and relatively PDS is to the effect of mouse bone marrow cells hemopoietic progenitor cell (being called for short CFU-E, BFU-E) and grain monosystem CFU-GM (being called for short CEU-GM).
Materials and methods
1, animal: the NIH mice, male, body weight 18-22 gram.
2, PDS presses the preparation of embodiment 1 method.
3, CFU-E, BFU-E, CFU-GM cultivate by Tang's pendant string " hematopoietic cell culture technique ".
4, PDS stimulated in vitro test: the PDS working solution is added in mouse bone marrow cells hemopoietic progenitor cell CFU-E, BFU-E, the CFU-GM cultivating system, and final concentration is respectively 1,5,10,20,50 μ g/ml, sets up matched group, cultivates observed result after 7 days.
5, the experimental result t method of inspection is analyzed the difference between PDS group and the matched group.
The result:
(1) PDS is to the effect of mouse bone marrow cells CFU-E
Table 1 shows the situation that different PDS is added 8 parts of mouse bone marrow cells cultivation CFU-E, the result show when PDS concentration be 1,5,10 and CFU-E colony productive rate is apparently higher than matched group (p＜0.05) during 20ug/ml, the raising rate is respectively 26.1%, 58.5%, 29.0% and 25.8%.
Table 1 PDS is to the effect of mouse bone marrow cells CFU-E
| Concentration (ug/ml) | CFU-E/0.5×10 5 | Raising rate (%) | The P value |
| 0 1 5 10 20 50 | 38.6±8.4 48.5±8.8 61.3±10.2 49.8±8.8 48.6±9.3 43.8±7.1 | - 26.1±3.3 58.5±6.1 29.0±2.2 25.8±3.0 13.5±2.1 | - ＜0.05 ＜0.01 ＜0.05 ＜0.05 ＞0.05 |
(2) PDS is to the effect of mouse bone marrow cells BFU-E
Table 2 shows that the PDS with variable concentrations adds 8 parts of mouse bone marrow cells cultivation BFU-E situations, the result shows that BFU-E colony productive rate is apparently higher than matched group (p＜0.05) when PDS concentration is 1,5,10 and 20 ug/ml, the raising rate is respectively 27.0%, 64.7%, 36.7% and 29.4%, and high concentration does not suppress.
Table 2 PDS is to the effect of mouse bone marrow cells BFU-E
| Concentration (ug/ml) | BFU-E/0.5×10 5 | Raising rate (%) | The P value |
| 0 1 5 10 20 50 | 8.5±2.5 10.8±2.6 14.0±5.9 11.6±2.7 11.0±2.7 9.8±2.4 | - 27.0±3.1 64.7±5.8 36.7±4.1 29.4±2.4 15.3±1.9 | - ＜0.05 ＜0.01 ＜0.01 ＜0.05 ＞0.05 |
(3) PDS is to the effect of mouse bone marrow cells CFU-GM
Table 3 shows that the PDS with variable concentrations adds 8 parts of mouse bone marrow cells cultivation CFU-GM situations, the result shows when PDS concentration is 1,5,10,20 and 50 ug/ml, CFU-GM colony productive rate is apparently higher than matched group (p＜0.05), and the raising rate is respectively 32.3%, 54.0%, 38.2%, 36.5% and 28.6%.
Table 3 PDS is to the effect of mouse bone marrow cells CFU-GM
| Concentration (ug/ml) | CFU-GM/2×10 5 | Raising rate (%) | The P value |
| 0 1 5 10 20 50 | 49.8±8.4 65.9±10.3 76.7±10.4 68.8±9.8 68.0±8.5 63.8±9.8 | - 32.3±3.1 54.0±6.1 38.2±4.1 36.5±3.5 28.6±2.6 | - ＜0.05 ＜0.01 ＜0.01 ＜0.05 ＜0.05 |
This experimental result shows: PDS is external all to have the effect of significantly stimulating proliferation to mouse bone marrow cells CFU-E, BFU-E, CFU-GM, compare with matched group and improve 58.5%, 64.7% and 54.0% respectively, and high concentration does not suppress.
Embodiment 3 panoxadiol are to the effect of normal person's bone marrow hematogenesis CFU-GM
Materials and methods
1, specimen: get the normal rib bone marrow of non-hematopathy patient, bone marrow smear is that normal marrow resembles.
2, CFU-E (CFU-E, BFU-E), grain single CFU-GM (CFU-GM) cultural method is identical with embodiment 1.
3, multipotential progenitor cells (CFU-Mix) is cultivated: get in the IMDM Mixed culture system and contain 4 * 10
5/ ml BMNC, 0.3% agar, 25% people AB type serum, 10
-5Mol/L 2-ME and L-glutaminate, the hemopoietic growth factor that every ml cultivating system contains reorganization is as follows: erythropoietin (EPO) 2U, grain/list-colony stimulating factor (GM-CSF) 20ng, be seeded on the culture plate with 0.25 ml/ hole (four multiple holes), place 37 ℃, contain 5%CO
2The incubator of saturated humidity is hatched, counting CFU-Mix colony after 14 days.
4, PDS is to CFU-E, BFU-E, CFU-GM and CFU-Mix irritant test: the PDS working solution is added in the cultivating system, final concentration is respectively 2.5,10,20,50,100 and during 200ug/ml, and set up the matched group of no PDS, observed result after cultivating 7,14 days.
5, data statistic analysis is handled and is checked with t.
The result:
(1) PDS is to the effect of normal person's bone marrow CFU-E
Table 1 shows that the PDS with variable concentrations adds the result that 10 parts of normal person's bone marrow prepares are cultivated CFU-E.When PDS concentration was 2.5,10,20,50,100 and 200 ug/ml, CFU-E colony productive rate had significant difference apparently higher than matched group.The raising rate is respectively 24.8%, 25.3%, 33.9%, 54.9%, 44.0%, 32.4%.
Table 1 PDS is to the proliferation function (n=10) of normal person's bone marrow CFU-E
| Concentration (ug/ml) | CFU-E/0.5×10 5 | Raising rate (%) | The P value |
| 0.0 2.5 10.0 20.0 50.0 100.0 200.0 | 115.2±20.6 143.8±20.8 144.3±29.6 154.3±28.2 178.5±30.2 165.9±22.4 152.5±25.2 | - 24.8±4.1 25.3±3.9 33.9±5.1 54.9±6.3 44.0±3.2 32.4±5.0 | - ＜0.05 ＜0.05 ＜0.01 ＜0.01 ＜0.01 ＜0.05 |
(2) PDS is to the proliferation function of normal person's bone marrow BFU-E
Table 2 shows the PDS of variable concentrations, adds 10 parts of normal person's bone marrow prepares cultivation BFU-E results.When PDS concentration was 2.5,10,20,50,100 and 200 ug/ml, BFU-E colony productive rate had significant difference all apparently higher than matched group.The raising rate is respectively 26.5%, 28.1%, 30.4%, 48.7%, 42.3% and 35.6%.Table 2 PDS is to the proliferation function (n=10) of normal person's bone marrow BFU-E
| Concentration (ug/ml) | BFU-E/0.5×10 5 | Raising rate (%) | The P value |
| 0.0 2.5 10.0 20.0 50.0 100.0 200.0 | 21.5±2.8 27.2±3.1 27.5±2.9 28.0±2.5 32.0±3.0 30.7±2.7 29.2±2.4 | - 26.5±3.1 28.1±4.2 30.4±4.1 48.7±5.1 42.3±4.6 35.6±4.1 | - ＜0.05 ＜0.05 ＜0.01 ＜0.01 ＜0.01 ＜0.01 |
(3) PDS is to the proliferation function (n=10) of normal person's bone marrow CFUGM
Table 3 shows that the PDS with variable concentrations adds 10 parts of normal person's bone marrow prepares cultivation CFU-GM results.When PDS concentration was 10,20,50,100 and 200 ug/ml, CFU-GM colony productive rate had significant difference all apparently higher than matched group.The raising rate is respectively 15.2%, 25.9%, 27.6%, 27.3% and 27.1%.
Table 3 PDS is to the proliferation function (n=10) of normal person's bone marrow cfu-gm
| Concentration (ug/ml) | CFU-GM/2×10 5 | Raising rate (%) | The P value |
| 0.0 2.5 10.0 20.0 50.0 100.0 200.0 | 165.9±18.6 173.1±19.5 182.8±22.1 208.7±23.4 211.7±26.3 211.2±25.9 210.8±22.9 | - 4.3±2.1 15.2±8.5 25.9±4.1 27.6±4.2 27.3±3.9 27.1±3.8 | - ＞0.05 ＜0.05 ＜0.01 ＜0.01 ＜0.01 ＜0.01 |
(4) PDS is to the proliferation function of normal person's bone marrow CFU-Mix
Table 4 shows that the PDS with variable concentrations adds 8 parts of normal person's bone marrow prepares cultivation CFU-Mix results.When PDS concentration was 10,20,50 ug/ml, CFU-Mix colony productive rate had significant difference apparently higher than matched group.The raising rate is respectively 22.4%, 48.9% and 31.6%.
Table 4 PDS is to the proliferation function (n=8) of normal person's bone marrow CFU-Mix
| Concentration (ug/ml) | CFU-Mix/1×10 5 | Raising rate (%) | The P value |
| 0.0 2.5 10.0 20.0 50.0 100.0 200.0 | 9.8±1.8 11.3±2.0 12.0±1.9 14.6±2.0 12.9±1.6 11.2±1.7 11.0±1.4 | - 15.3±1.7 22.4±1.9 48.9±3.9 31.6±2.8 14.3±1.6 12.2±1.3 | - ＞0.05 ＜0.05 ＜0.01 ＜0.01 ＞0.01 ＞0.01 |
This result of study shows: 1, PDS is that the hemopoietic progenitor cell of three kinds of series all has the effect of significantly stimulating proliferation to the red system of normal person's bone marrow, grain system, macronucleus, and this multiplication effect is similar to hemopoietic growth factor.Can not improve the colony productive rate even increase this somatomedin concentration again when 2, cultivating hemopoietic progenitor cell according to the hemopoietic growth factor of former result of study applying high density, PDS then can play synergism and improve the colony productive rate significantly with somatomedin.This synergism clinically may be more meaningful, because most hematopathy patient's body is interior and have no lack of hemopoietic growth factor, the patient who has also is higher than the normal person.3, the PDS of 10-50ug/ml concentration is the appropriate concentration of stimulated in vitro hemopoietic progenitor cell propagation, also can be the clinical trial dosage simultaneously reference is provided.
Embodiment 4 Ginsengdiol histsaponins are to the in-vitro multiplication effect of Patients with Aplastic Anemia hemopoietic progenitor cell
Case: 30 examples are diagnosed as aplastic anemia (AA) patient, and Chinese Journal of Hematology is pressed in diagnostic criteria, 1987:8 (8) back cover and 463 " aplastic anemia diagnosis and criterion of therapeutical effect ".Wherein male 17 examples, women 13 examples, The median age 32 years old (7-66), acute aplastic anemia (AAA) 8 examples, chronic aplastic anemia (CAA) 22 examples.Again by CFU-GM colony number, the peripheral blood mononuclear cell of patient (PBMNC) is divided into androgen response type (AA1) 15 examples to the inhibition test of normal CFU GM and CFU-E to the sensitivity of first testis, and immune-mediated type (AA2) 8 examples and stem cell lack three types of type (AA3) 7 examples.
Medicine: PDS extracts routinely, is made into 1 mg/ml working solution with culture medium, positive press filtration, and 4 ℃ of preservations are standby.
Hemopoietic progenitor cell culture method: PDS is that final concentration adds in the aplastic anemia patient bone marrow In vitro culture system with 25 μ g/ml, and sets up the matched group that does not add PDS.
The result: (1) PDS is to the effect of androgen response type aplastic anemia (AA1) 15 routine bone marrow CFU-E, BFU-E and CFU-GM, the result shows PDS group CFU-E, BFU-E and CFU-GM colony productive rate all apparently higher than matched group (p＜0.05), and the raising rate is respectively 38.1%, 47.6% and 23.8% (seeing Table 1).
Table 1
| The colony value | Matched group | PDS | Raising rate (%) | The P value |
| CFU-E BFU-E CFU-GM | 31.0±5.4 2.1±0.6 54.3±8.5 | 42.8±7.2 3.1±0.7 67.2±11.0 | 38.1±4.2 47.6±4.3 23.8±2.1 | ＜0.01 ＜0.01 ＜0.05 |
(2) PDS is to the effect of immune-mediated type aplastic anemia (AA2) 8 routine bone marrow CFU-E, BFU-E and CFU-GM, the result shows: PDS group CFU-E, BFU-E compare no significant difference (p＞0.05) with CFU-GM colony productive rate with matched group, the raising rate is respectively 13.3%, 11.1% and 12.9% (seeing Table 2).
Table 2
| The colony value | Matched group | PDS | Raising rate (%) | The P value |
| CFU-E BFU-E CFU-GM | 10.5±3.4 0.9±0.4 23.2±5.1 | 11.9±3.6 1.0±0.4 26.2±5.8 | 13.3±1.4 11.1±1.1 12.9±1.3 | ＞0.05 ＞0.05 ＞0.05 |
(3) PDS lacks the effect of type aplastic anemia (AA3) 7 routine bone marrow CFU-E, BFU-E and CFU-GM to stem cell, the result shows: PDS group CFU-E, BFU-E compare no significant difference (p＞0.05) with CFU-GM colony productive rate with matched group, the raising rate is respectively 3.8%, 0% and 6.0% (seeing Table 3).
Table 3
| The colony value | Matched group | PDS | Raising rate (%) | The P value |
| CFU-E BFU-E CFU-GM | 2.6±0.9 0 5.0±1.3 | 2.7±1.0 0 5.3±1.5 | 3.8±0.4 0 6.0±0.5 | ＞0.05 - ＞0.05 |
This experimental result shows: PDS is to the effect that also stimulates proliferation of aplastic anemia patient's hemopoietic progenitor cell.And immune-mediated type is because of existing the inhibition system, stem cell lack type because of CFU-GM very little, the effect of PDS can not be expressed.When its when sb.'s illness took a favorable turn, it is responsive that the PDS in vitro tests also tends to.
Embodiment 5 Ginsengdiol histsaponins and compositions thereof are to intractable hemopathic observation of curative effect
Case: chronic aplastic anemia anemia (CAA) 23 examples, wherein male 11 examples, women 12 examples, age 11-72 year, middle several sick times 6 (1-13) year.The past medicine comprises: Chinese medicine, androgen, adrenal gland's glucocorticosteroid hormone, HGF, cyclosporin A and Supporting Therapys such as blood transfusion and antibiotic.17 examples through more than fail to respond to any medical treatment, in addition 6 routine hemoglobin and leukocyte have rising, but that platelet still is in is low-level.
Thrombocytopenic purpura (ITP): 33 examples, the course of disease 2.5 (1-8) year.Adult group man 9 examples, women 19 examples, middle ages 43 several years (22-71) years.The child organizes 5 examples and is the male, year at age 8 (6-13).Most of patients all responds through the glucocorticoid treatment initial stage, and is later invalid.Child's case all through prednisone, vincristine and azathioprine etc. separately or therapeutic alliance all invalid.
Therapeutic outcome: (1) aplastic anemia phases 17 example, treatment effective percentage 70.6% (12/17).Average platelet 31.6 * 10 before the treatment
9/ L, hemoglobin 51.3g/L and leukocyte 2.8 * 10
9/ L; Bring up to 64.7 * 10 after the Ginsengdiol histsaponin treatment
9/ L, 90.3g/L and 4.06 * 10
9/ L has significant difference (p＜0.01), and platelet, hemoglobin and leukocytic raising rate are respectively 144.0%, 20.1% and 36.3%.Wherein cure 5 examples, alleviation 4 examples, progressive 3 examples, invalid 5 examples (keep by blood transfusion before the 2 example treatments in 5 examples, do not transfuse blood in a year after the treatment again, hemogram has and rises, but does not arrive " progress " standard) substantially.
(2) aplastic anemia anemia convalescent period (aplastic anemia treatment back hemoglobin and leukocyte have rising, but the still low person of platelet) 6 examples, this group effective percentage is 100% (6/6).Wherein cure 4 examples, alleviation 1 example, progressive 1 example substantially.4 examples platelet after the Ginsengdiol histsaponin treatment reaches normally, and the also more former level of hemoglobin and leukocyte raises, and 1 routine platelet is by treatment preceding 15.0 * 10
9/ L rises to 96.0 * 10
9/ L.
(3) thrombocytopenic purpura 33 examples, the treatment effective percentage is 84.8% (28/33).
Adult group: 28 examples, average platelet is 36.2 * 10 before the treatment
9/ L brings up to 82.4 * 10 after the Ginsengdiol histsaponin treatment
9/ L, the raising rate is 127%, credit is analysed and is had significant difference (p＜0.01) by statistics.Wherein basic cure and produce effects 10 examples, very imitate 9 examples, 7 examples, invalid 2 examples improve, effective percentage is 92.8% (26/28), 1 example is simultaneously with the HbsAg positive in the produce effects, after Ginsengdiol histsaponin is treated 8 months, hepatitis B three be turn out cloudy, platelet reaches normal, follow up a case by regular visits to not recurrence in 2 years 10 months, meet criterion of cure.
Child's group: 5 examples, curing effective percentage is 40% (2/5), wherein produce effects and good each 1 example, invalid 3 examples of imitating.
Aplastic anemia of being brought out by hepatitis B in (4) 56 examples and thrombocytopenic purpura 17 examples, curing effective percentage is 88.2% (15/17).Positive or other antibody positives of HbsAg are all arranged, and wherein aplastic anemia 7 examples are treated through Ginsengdiol histsaponin: wherein cure 2 examples, progressive 3 examples, invalid 2 examples substantially, effective percentage 71.4% (5/7); Thrombocytopenic purpura 10 examples, treat through Ginsengdiol histsaponin: cure and produce effects 6 examples, very imitate 3 examples, 1 example that improves wherein substantially, effective percentage reaches 100%.2 examples are arranged in 17 examples after the Ginsengdiol histsaponin treatment, HbsAg transfers feminine gender to from the positive.
This result of study shows: Ginsengdiol histsaponin has the class somatomedin effect of hemopoietic progenitor cell proliferation.Clinical practice obtains gratifying curative effect, and total effective rate is 82.1% (46/56).Interior curative effect is consistent with the experiment in vitro result.CAA and adult ITP are frequently-occurring disease and refractory disease, cause of disease complexity in hematopathy.Ginsengdiol histsaponin can make CAA and ITP peripheral blood obviously rise, and raises obviously with platelet especially.28 example adult ITP treat through Ginsengdiol histsaponin, and effective percentage is 98.2%.ITP is an autoimmune disease, and ginseng diol saponin may increase normal immunity or suppress autoantibody.CAA that 17 routine hepatitis ies cause and ITP have positive or other antibody positives of HbsAg, and 15 examples obtain good efficacy, and wherein 2 routine HbsAg are by turning out cloudy property of the positive.The prompting panoxadiol has good effect to immune system.
Embodiment 6 Ginsengdiol histsaponins are to people CD34
+The effect of hematopoietic stem
Material and method
1. getting surgical operation does not have blood patient's rib, separates with conventional method, obtains BMNC.
2. get mononuclearcell and obtain CD34 with immunomagnetic beads labeling of monoclonal antibody method
+Cell, through flow cytometry analysis purity up to more than 90%.
3.PDS extracting method identical with embodiment 1.
4. hemopoietic progenitor cell is cultivated: CD34
+Cell 1 * 10
4/ ml, cultivating system contain interleukin-3 (IL-3), GM-CSF, EPO.37 ℃ of 5%CO
2Cultivated 14 days.Observe BFU-E, CFU-E, CFU-E, CFU-E and CFU-Mix colony number.Not add the PDS group is contrast.The results are shown in Table 7 table 7 PDS to CD34
+The proliferation function of hematopoietic stem (X+SD, n=5)
| PDS (μg/ml) | BFU-E | Raising rate (%) | CFU-E | Raising rate (%) | CFU-GM | Raising rate (%) | CFU-Mix | Raising rate (%) |
| 0 5 10 20 50 100 | 28.6±2.1 56.0±4.3 70.1±7.0 78.0±7.8 68.1±5.9 43.1±4.4 | 95.8±8.0 145.0±10.4 172.0±12.1 186.0±10.1 50.1±8.7 | 69.0±3.1 110.0±5.0 128.3±6.8 154.0±8.1 146.0±7.9 112.0±5.6 | 59.4±4.1 85.9±7.1 123.1±11.4 111.1±9.8 62.3±5.6 | 56.0±2.1 62.0±4.2 68.4±5.5 83.1±9.1 140.6±8.8 98.2±7.6 | 10.1±0.9 21.4±1.8 48.2±5.3 151.0±11.2 75.0±8.7 | 28.1±1.8 47.2±2.2 49.3±3.1 64.0±5.5 60.2±5.9 54.0±6.0 | 67.2±8.3 75.4±5.9 127.0±1.8 113.5±9.8 92.1±7.8 |
Research in the past only can be at the hemopoietic progenitor cell than late period.And along with the isolation and purification method of hematopoietic stem cell makes progress, CD34
+Hematopoietic stem is the hematopoietic cell of the present earliest period that can study, has demonstrated huge clinical value, CD34
+Hematopoietic stem is transplanted the method that has become radical cure malignant hematologic disease and some solid tumor, but the reconstitute hematopoiesis function also can be used for heredopathia and genetic treatment of tumor.The proliferation function of PDS to hematopoietic stem observed in this research.The result shows that PDS not only stimulates the hemopoietic progenitor cell propagation than late period, and to CD34
+Cell forms all kinds of directed hemopoietic progenitor cell colonies and has tangible effect.PDS promotes CD34
+Cell forms BFU-E, CFU-E, CFU-GM and CFU-Mix colony number and increases by 186.0 ± 10.1%, 123.1 ± 11.4%, 151.0 ± 11.2% and 127.0 ± 1.8% respectively, and the raising rate is significantly higher than PDS to the hemopoietic progenitor cell than late period.Prompting PDS not only has the CD34+ of promotion cell proliferation, and can induce the two-way function of its directed differentiation, and to early stage more hematopoietic cell, acts on obvious more.The patient of these results suggest after for bone marrow transplantation and chemotherapy uses the PDS treatment can play synergism with hemopoietic growth factor, to improving curative effect and recovering hemopoietic will be effectively, for intractable hematopathy, pathological changes then can be by promoting early stage CD34 ancestral cells stage person
+Hematopoietic stem is bred and is broken up and the recovery hemopoietic function.
The PDS concentration of 5-50 μ g/ml is to stimulate CD34
+The optium concentration that cell colony forms when concentration not only occurs suppressing phenomenon during up to 100 μ g/ml, and still can improve the colony number, illustrates that also PDS quality and purity are good.
The Study on mechanism of embodiment 7 panoxadiol's hemopoietic progenitor cell proliferations
How PDS impels the mechanism of action of hemopoietic progenitor cell propagation not understand as yet.Transcription factor c-Fos, c-Jun and GATA-2 play an important role in hemopoietic, and their major function is the reaction of mediation hematopoietic cell to somatomedin, participates in the regulation and control of cell proliferation and differentiation related gene.Because the effect of PDS hemopoietic is similar to somatomedin,, inquire into the signal pipeline of PDS in hematopoietic cell so use for reference the method for research somatomedin.
Material and method
1, human cell line cultivate and the PDS stimulation process to select people's grain for use be that cell strain HL-60, erythroid cells strain K562, macronucleus are that cell strain CHRF-288 and Meg-01 do target cell.The conventional cultivation in containing the IMDM culture medium of 10% calf serum, be divided into 2 groups: experimental group adds PDS 5ug/ml respectively, matched group does not add PDS, cultivated 1 month, cell is done hungry the processing, and serum deprivation was hatched 24 hours, added PDS 25 ug/ml then respectively, cell and drug effect extracted nucleoprotein after 2 hours.
2, extract nucleoprotein nucleoprotein extracting method being improved by the Dinam report, cell is after cold PBS washing, be suspended in the buffer A, contain protease inhibitor: aprotinin (Leupeptin), papain peptide for inhibiting (antipain), bright peptide for inhibiting (aprotinin) and pepsin inhibitor (pepstatin) are (Sigma), be 10ug/ml concentration, broken cell membrane after vibrating, collecting cell nuclear, be suspended among the buffer C, ultrasonic disintegrator smudge cells nuclear, centrifugation 12000 rpm, 20 minutes, get supernatant, measure protein concentration.
3, after protein immunoblot analysis (Western Blot) 10ug nucleoprotein adding sds gel sample loading buffer 25ul boils, after the SDS-PAGE gel electrophoresis separates.The electricity consumption of albumen galley proof is transferred on the nitrocellulose, add specific anti-human c-Fos, c-Jun and GATA-2 antibody (Santa Cruz) 0.5ug/ml, room temperature association reaction 1 hour, add two anti-(dilutions in 1: 2000) of horseradish peroxidase-labeled, ECL luminous agent (Amersham) effect 1 minute, exposed 1-10 minute, the back optical density scanner analysis result that develops, same experiment repeats three times.PDS examines interior c-Fos, c-Jun and GATA-2 transcription factor content after handling cell as a result
1.c-Fos:HL-60, K562, CHRF-288 and Meg-01 four strain cells are after the PDS stimulation process, Western Bolt shows nucleoprotein and c-Fos antibodies reaction zone, after the densitometric scan analyzing and processing, it is 1.5-2.0 times that the C-FOS transcription factor content of four strain cells is higher than undressed cell.
2.c-Jun:HL-60, K562, CHRF-288 and Meg-01 four strains, HL-60 cellular expression c-Jun, and no change before and after the PDS stimulation process.And K562, CHRF-288 and Meg-01 three strain cellular expressions are very low, and PDS handles the back obviously to be increased, and after the densitometric scan analyzing and processing, the c-Jun transcription factor content of three strain cells is higher than undressed cell, are 2.0-3.0 times.
3.GATA-2:HL-60, K562, CHRF-288 and Meg-01 four strain cells are after the PDS stimulation process, Western Bolt shows nucleoprotein and GATA-2 antibodies reaction zone, and the GATA-2 transcription factor content of CHRF-288 and Meg-01 two strain cells is undressed cell 1.4-1.8 times.
But the change of somatomedin, cytokine and other extracellular signaling molecule inducing cell propagation, differentiation and function and behavior.This cell has important biological significance to the reaction of external environment, need signal transmission and gene regulation program through a series of complexity, comprise combining of signaling molecule and surface receptor and part, and then trigger signal pipeline in the specific cell, and the promotor gene transcript and expression.Therefore, the interior signal pipeline of cell is as the bridge that links genes of interest in cell surface and the nucleus.
After the PDS stimulation process, the c-Fos protein content of HL-60, K562, CHRF-288 and Meg-01 cell is higher than unprocessed cell.The c-Jun of K562, CHRF-288 and Meg-01 cell expresses obviously to be increased.And GATA-2 only is higher than unprocessed cell at megalokaryocyte CHRF-288 and the Meg-01 protein content that PDS handles.These results suggest PDS is the hemopoietic cell proliferation by inducing c-Fos, c-Jun and GATA-2 transcriptional regulation protein respectively, from the class somatomedin effect of molecular level proof PDS.Red system, grain system and macronucleus are that transcriptional regulation protein itself is expressed the different of height in three kinds of serial nucleus, and to the sensitivity level difference of PDS, this is because due to the serial specificity.
Claims (2)
1, the application of compositions in treating hematopathy of Ginsengdiol histsaponin and pharmaceutical carrier and/or pharmaceutical excipient composition.
2, compositions as claimed in claim 1, the especially application in intractable treating hematopathy.
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101579376B (en) * | 2008-05-14 | 2012-01-25 | 北京卓越同创药物研究院 | Diol saponin traditional Chinese medicine preparation and preparation method thereof |
| EP2517711A4 (en) * | 2009-12-21 | 2013-05-29 | Lion Corp | Hyperlipemia-ameliorating agent, anemia-ameliorating composition, uric-acid-level-reducing composition, and foods and beverages |
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1999
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Cited By (8)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101579376B (en) * | 2008-05-14 | 2012-01-25 | 北京卓越同创药物研究院 | Diol saponin traditional Chinese medicine preparation and preparation method thereof |
| EP2517711A4 (en) * | 2009-12-21 | 2013-05-29 | Lion Corp | Hyperlipemia-ameliorating agent, anemia-ameliorating composition, uric-acid-level-reducing composition, and foods and beverages |
| CN103520179A (en) * | 2009-12-21 | 2014-01-22 | 狮王株式会社 | Hyperlipemia-ameliorating agent, anemia-ameliorating composition, uric-acid-level-reducing composition, and foods and beverages |
| EP2702996A1 (en) * | 2009-12-21 | 2014-03-05 | Lion Corporation | Anemia-ameliorating composition |
| US8927033B2 (en) | 2009-12-21 | 2015-01-06 | Lion Corporation | Hyperlipemia-ameliorating agent, anemia-ameliorating composition, uric-acid-level-reducing composition, and food or beverage |
| CN103520179B (en) * | 2009-12-21 | 2015-09-09 | 狮王株式会社 | Hyperlipidemia improving agent, anemia improve compositions, uric acid level reduces compositions and beverage/food |
| KR20160101724A (en) * | 2009-12-21 | 2016-08-25 | 라이온 가부시키가이샤 | Hyperlipemia-ameliorating agent, anemia-ameliorating composition, uric-acid-level-reducing composition, and foods and beverages |
| KR101717893B1 (en) * | 2009-12-21 | 2017-03-17 | 라이온 가부시키가이샤 | Hyperlipemia-ameliorating agent, anemia-ameliorating composition, uric-acid-level-reducing composition, and foods and beverages |
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| CN1128621C (en) | 2003-11-26 |
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