In recent years, there has been a global boom in stem cell research, and the efficacy of stem cell therapy is gradually being recognized. Similarly, stem cell therapy has been widely used in the field of hepatobiliary and pancreatic diseases. In addition to traditional drug therapy and surgery, stem cell therapy will undoubtedly be another important option for hepatobiliary and pancreatic diseases. In this paper, we intend to discuss the application of stem cell therapy in the clinical treatment of hepatobiliary and pancreatic diseases.
The existence of stem cells in hepatobiliary and pancreatic organs has been confirmed, which provides a theoretical basis for stem cell therapy in hepatobiliary and pancreatic diseases. In the liver, hepatic oval cells have been shown to be involved in liver regeneration as hepatic stem cells. In the pancreas, Seaberg et al [1] suggested that pancreatic stem cells are present not only in the pancreatic ducts, but also in the islets themselves.
Also in malignant tumors of the liver, gallbladder and pancreas, there are more studies confirming the presence of stem cells in hepatocellular carcinoma, gallbladder cancer and pancreatic cancer. Some cell surface molecular markers are thought to be markers of stem cells in hepatobiliary and pancreatic cancers. In hepatocellular carcinoma CD133 [2, 3], EpCAM [4, 5] and CD90 [6] are thought to be associated with hepatocellular carcinoma stem cells. In pancreatic cancer studies have found that a subpopulation of CD44+/CD24+/ESA+ cells in pancreatic cancer cell lines is the most tumorigenic and is considered to be the stem cell population in pancreatic cancer [7]. There are also more studies that suggest that CD133 is also one of the surface markers of pancreatic cancer stem cells. A subpopulation of CD44+/CD133+ cells was found to have stem cell properties in gallbladder cancer cell lines [8, 9]. Many other studies have confirmed the presence of side population cells in hepatobiliary and pancreatic cancer cells, which is also the basis for the presence of cancer stem cells.
The hepatobiliary and pancreatic glands are the main digestive organs of the human body, which have relatively high cellular metabolism and are therefore more prone to cellular damage and degenerative diseases. Stem cells have the characteristics of long-term survival, continuous self-propagation, cross-lineage and cross-germline differentiation, strong plasticity, chemotaxis, etc. Therefore, stem cell therapy is applicable to more diseases of hepatobiliary and pancreatic.
Stem cells used in hepatobiliary and pancreatic diseases are divided into two categories: embryonic stem cells and adult stem cells. Embryonic stem cells are mainly umbilical cord blood stem cells and umbilical cord stem cells, which have the advantages of strong cell proliferation ability, but have the disadvantages of tumorigenicity after transplantation, cannot be autologous transplantation, and have certain ethical issues. The advantages of adult stem cells are no rejection reaction, no tumorigenicity, no ethical problems, etc., but the disadvantage is that the cell proliferation ability is not as strong as embryonic stem cells.
1.Liver diseases.
The indications are mainly for post-hepatitis cirrhosis decompensated stage, acute, subacute and chronic heavy hepatitis, drug-related liver disease, etc. Studies in the Department of Gastroenterology of Southwest Hospital concluded that stem cell therapy is not effective in alcoholic cirrhosis, and the specific mechanism is unknown. The main route of input is via interventional input of stem cells into the hepatic artery.
Both domestic and international studies have found [10-13] that autologous bone marrow stem cell transplantation for end-stage liver disease generally results in significant changes in laboratory parameters at week 2. It was concluded that serum albumin levels increased significantly in all patients at week 4 after transplantation [13], and there were also results showing a significant increase in albumin levels after 8 [11] or 24 [10] weeks after transplantation, which were the most significant and long-lasting changes in laboratory indicators. ALT, AST and Child scores also decreased significantly at week 4 postoperatively.
However, serum total bilirubin levels decreased without statistical significance, and only one study concluded that total bilirubin decreased significantly after 8 weeks [11]. SHUJI [10] in Japan even compared immunohistochemical AFP and PCNA staining of preoperative and postoperative liver puncture pathology in three cases in their study and found that both AFP and PCNA were significantly enhanced at 4 weeks after stem cell treatment. The above results indicated that the liver function of patients significantly improved after bone marrow stem cell transplantation treatment, especially the synthetic and repair function of the liver was significantly improved. These changes, especially the elevation of serum albumin, were impossible to achieve with general liver protection therapy. In addition, we observed a significant improvement in subjective clinical symptoms, such as bloating, poor appetite and weakness, and a significant improvement in quality of life in most patients 2 weeks after transplantation. No serious adverse reactions and complications were found in all transplant patients.
The Department of Hepatobiliary Surgery of the Third Affiliated Hospital of Sun Yat-sen University in China reported for the first time [14] that intraoperative “portal vein catheter-subcutaneous drug box” was embedded in the surgical treatment of portal hypertension in cirrhosis, and the patients in the treatment group were infused with autologous bone marrow cells through the transplantation channel 3-4 weeks after surgery. After the stem cell treatment, the albumin, transaminase, coagulation, total bilirubin and liver fibrosis indexes of liver function were significantly improved compared with those before the stem cell treatment.
However, we believe that the method of extracting autologous bone marrow stem cells in this study was crude, and the total number of individual nucleated cells obtained was low, only (8.6±3.7)×10 7, which failed to reach the current more recognized cell volume of 109. In addition, the study introduced stem cells into the liver through the portal vein, whereas patients with portal hypertension in cirrhosis often have portal blood flow against the liver, and stem cells may not flow into the liver through the portal blood flow. Moreover, the intraoperative burial of the subcutaneous cassette is more traumatic, and there is a possibility of vascular hemorrhage after exclusion. Therefore, the study has some defects. In 2010, the Department of Hepatobiliary Surgery of our hospital (The First Hospital of Wenzhou Medical College) started to find the right gastric artery in the surgical treatment of portal hypertension, and after entering the hepatic artery through the right gastric artery via pediatric deep vein placement, the autologous bone marrow stem cells were obtained by negative collection method, and the amount of individual nucleated cells reached 109 or more. Five cases have been treated and all of them have achieved good results. The albumin, transaminases and coagulation of liver function of patients after stem cell treatment have improved significantly compared with the previous ones.
It can be said that stem cell transplantation for liver disease is currently the most studied, the most well-documented and the most definite effect considered among all clinical disease applications. We think this may be related to the stronger ability of stem cells to promote liver regeneration and the better objective evaluation index of liver function improvement.
2.Pancreatic diseases.
1.Diabetes mellitus
The new study concluded that the sharp decrease of pancreatic β-cells in the pancreas and the defect of insulin secretion function are one of the main reasons for the occurrence of diabetes mellitus. Therefore, how to stop the destruction of β cells and promote the re-growth of β cells is an important measure for the treatment of diabetes.
In type 1 diabetes, the Journal of the American Medical Association (JAMA) published in 2007 a study by Brazilian researcher Ju lio C. Vo ltarelli [15] on the short-term reconstitution of endogenous insulin secretion from β cells in T1DM patients with newly diagnosed T1DM by autologous bone marrow mesenchymal stem cell transplantation technique. In this study, 15 patients with T1DM were enrolled, and 14 patients were taken off insulin therapy after treatment, including one patient who was off insulin therapy for 35 months and the two shortest cases who were off insulin therapy for 1 and 5 months, respectively. After treatment, the mean area under the C-peptide curve improved significantly compared with that before treatment and remained stable for 24 months. The preliminary findings suggest that endogenous insulin secretion can be re-established in newly diagnosed T1DM patients by autologous HSCT technique, inducing a significant period of time off insulin therapy in most patients with T1DM. C-peptide levels were significantly elevated after transplantation, and most patients were successfully freed from insulin dependence.
For type 2 diabetes, in 2009, Bhansali et al [17], an Indian scholar, selected 10 patients with a history of type 2 diabetes (GAD antibody negative) for more than 5 years, all of whom had failed on triple oral medications and were currently on insulin doses >0. 7 U /kg /d for at least 1 year. After 6 months of autologous stem cell treatment, 7 out of 10 cases had a 75% reduction in insulin dosage compared to baseline, a 1% decrease in glycosylated hemoglobin (H bA1C), a >50% reduction in insulin dosage, and an increase in insulin and C-peptide secretion, and no serious side effects were found. In China, the Affiliated Hospital of Qingdao University reported [18] that 61 cases of T2DM were treated with autologous bone marrow stem cells transplanted via pancreatic vascular cannula, of which 49 cases showed significant efficacy (81.7% efficiency).
In general, it seems that the overall efficiency can reach 80%-90% for all types of diabetes mellitus, and most patients show a reduction in the original dose of oral medication and a reduction in the dose of insulin used; some patients can gradually reach discontinuation of insulin or even discontinuation of oral medication, and the efficacy is more obvious especially in relatively early patients; the duration of discontinuation of medication or the duration of reduction in the dose of medication is probably related to the time patients receive the treatment. The duration of treatment may be related to the time the patient receives.
2, pancreatitis
Chronic pancreatitis refers to persistent damage to pancreatic tissue and function caused by various etiologies, eventually leading to permanent loss of endocrine and exocrine function of the pancreas. More basic studies have confirmed that the degree of pancreatic lesions and fibrosis in rats with chronic pancreatitis treated with mesenchymal stem cells were significantly reduced [19-21]. A study from Tianjin Medical University showed [19] that connective tissue growth factor, transforming growth factor 13, collagen type I and IIJ levels and myeloperoxidase activity were significantly reduced in pancreatic tissue. It was suggested that bone marrow mesenchymal stem cells had a significant repair effect on pancreatic injury in rats with chronic pancreatitis, and the mechanism may be related to the inhibition of connective tissue growth factor and transforming growth factor 13 production, suppression of inflammatory response, and reduction of collagen proliferation.
Columbia University studied [20] chronic pancreatitis rats molded with frog skin and treated with bone marrow stem cell transplantation and found that bone marrow stem cells significantly promoted the regeneration of pancreatic stellate cells to repair pancreatic tissue, but did not develop pancreatic tumors. In China, Jiang Xueliang [21] et al. labeled autologous bone marrow mesenchymal stem cells (MSCs) with the nuclear dye Hoechst 33258 after causing a pancreatitis model and infused back into the autologous bone marrow cavity, and found that 2 weeks after modeling, all MSCs from surviving rats admixed into the injured pancreas had differentiated into cvtokeratin 19 staining-positive cells, thus confirming the involvement of autologous bone marrow MSCs in physiological renewal and pathological regeneration of the pancreas.
Stem cell transplantation for the treatment of chronic pancreatitis has been supported by basic research and has been reported sporadically in clinical practice, but has not yet been published in the formal literature.
Cui et al [22] treated L_arginine-induced SAP by transplantation of bone marrow mesenchymal stem cells or injection of granulocyte colony-stimulating factor, and the blood amylase levels measured at 48 h and 72 h in the treated group were significantly lower than those in the control group. Pathological examination showed that the degree of pancreatic damage in the treated group was also less than that in the control group, suggesting that bone marrow MSCs helped to reduce the condition of SAP. A recent Korean study [23] showed that the infusion of bone marrow MSCs into SD rats with acute pancreatitis through the tail vein significantly reduced cytokines in the blood and decreased the expression of CD3+ T cells and Foxp3+ in pancreatic tissue, indicating that bone marrow MSCs could improve the inflammatory response and regulate autoimmunity in SAP.
Probably considering economic factors, practicality and ethical issues, no clinical application of stem cell therapy for acute pancreatitis has been reported.
3.Hepatobiliary and pancreatic malignancies.
As mentioned earlier, the existence of stem cells in hepatobiliary and pancreatic malignancies has been confirmed, and targeted therapeutic strategies targeting hepatobiliary and pancreatic cancer stem cells will become a new direction in hepatobiliary and pancreatic cancer treatment, which has the advantages of strong therapeutic specificity, significant effect, and basically no damage to normal tissues. Currently, there are three broad categories of specific strategies: molecular targeting of molecules specific to the surface of cancer stem cells; strategies to differentiate cancer cells to normal cells; and monoclonal antibody therapy against cancer stem cells. For example, Lin et al [24] applied NSC74859 (a specific inhibitor of STAT3) in TGF-β inactivated hepatocellular carcinoma cells to significantly inhibit tumor growth, and although both CD133+ and CD133C cells were sensitive to NSC74859 with an IC50 of 100 μmol/L, the inhibitor significantly blocked tumor growth in vivo, thus, IL6 /STAT3 is expected to be a better therapeutic target for hepatocellular carcinoma.
A study from Sun Yat-sen University in China [25] showed that blocking the Hedgehog signaling pathway may become a stem cell-targeted therapy for pancreatic cancer. Currently, there is no clinical application of stem cell therapy for hepatobiliary and pancreatic malignancies, but we believe that the concept of cancer stem cells as a therapeutic target has gained popularity and will certainly lay a solid foundation for future breakthroughs in clinical treatment of tumors.
Review and prospect of cell isolation technology for clinical application: At present, the domestic stem cell isolation technology is still relatively confusing. Because stem cells applied to human body cannot be sorted by antibody labeling, most of them used centrifugation method to sort stem cells in the past. Because of the difference in size, morphology and density between stem cells and other cells, differential centrifugation method, density gradient centrifugation method and magnetic bead sorting method are used. However, most of the so-called “stem cells” sorted out are of poor quality. In addition, there are also stem cells obtained by in vitro culture and expansion of umbilical cord blood or umbilical cord mesenchymal stem cell isolation, but all of them have certain ethical barriers and safety hazards. At present, the better one is the stem cell isolation kit produced by Ningxia Zhonglianda Company applying the principle of negative collection.
This kit uses a combination of negative collection method and density method to remove LIN antigen-positive cells and non-monocytes, so that there is no marker on the surface of the remaining stem cells and no loss of stem cells, which is easy to operate and convenient for clinical application. We used this method to extract 150 ml of stem cells from bone marrow blood, and the percentage of single nucleated cells was over 80% and the percentage of CD34+ cells was about 0.3%. We expect a better stem cell isolation technique with easy and reliable clinical application to appear and be applied to clinical treatment of stem cells.
Problems and outlook: The clinical follow-up of stem cell therapy for hepatobiliary and pancreatic diseases is not long yet, and its safety and efficacy are yet to be observed in the follow-up. There are still many issues that need to be studied more carefully and deeply in stem cell therapy for hepatobiliary and pancreatic diseases. For example, what are the indications of stem cell therapy for specific diseases of hepatobiliary and pancreatic diseases? Is multiple stem cell transplantation still effective, and what is the optimal interval between multiple treatments? Should the number of transplanted stem cells be individualized depending on the patient’s elevation, weight, and organ volume? And so on.
In addition, stem cells combined with tissue engineering should be an important direction for stem cell therapy for hepatobiliary and pancreatic diseases in the future. This is another important breakthrough in medical development, as it is expected to make “artificial liver” transplants possible.
The 21st century is gradually transitioning from the era of “organ transplantation” to the era of “organ manufacturing” and from the era of “drug therapy” to the era of “cell therapy”. “It is believed that stem cell therapy for hepatobiliary and pancreatic diseases will achieve more and greater breakthroughs.