1. Introduction
The human microbiota or microbiota live symbiotically in our body. In our gastrointestinal tract approximately 1013-14 microorganisms (mainly bacteria) live This number is said to be equal to human cells [1] . The amount of genomic content is over 100 times as compared to the whole human genome. Since the gut microbiota is involved in many different metabolic activities the gut microbiota is also known as the second brain since an enteric nervous system communicates with the brain via the nervous system [2] .
Each compartment of the digestive tract has a diverse microbial population and functions since environmental conditions change in each compartment. The major environmental conditions are such as acidic nature of gastric juices, the presence of bile salts, pancreatic enzymes, presence of mucous layer of pathogenic microorganisms.
There are significant differences in concentration and diversity of microbial communities ranging 102 per gram in the stomach to 1014 per gram in the colon [3] [4]
2. Microbiota and Diseases
Tremendous amounts of data indicate that variations and changes in the composition of the gut microbiota contribute to diseases such as Alzheimer’s disease, Parkinson’s disease, autism, depression to obesity and diabetes mellitus [5] . It has been experimentally validated that the gut microbiota influences distant organs such as lung, heart, liver, kidney and the central nervous system.
In order to tackle this subject, Long Li et al. [6] developed Amadis, a manually curated database that microbiota disease associations experimentally supported.
Here, I discuss diseases of which we have been working these days.
2.1. Depression
We all suffer and grieve at some point in our life. It is an unescapable response and such response may be needed for us to overcome such unhappiness in the future. Anxiety may prepare us to deal with stress by sharpening your senses.
In life we often lose beloved persons or cannot achieve our objectives successfully. In such cases we grieve. This is called situational depression. But when your depression last long you feel sad or do not intend to work more.
This is not caused by affairs in the world outside but by the brain chemistry.
The disease affects up to 15% of the general population and accounts for 12.3% global disease burden [7] .
Weight fluctuation and insomnia accompany depression. Gut problems such as diarrhea or constipation are often involved either as cause or effects.
Sometimes depressive symptoms are experienced as a comorbidity of other diseases. When we are sick we feel depressed and lose hope for the present or future life. Almost every disease increases our chance of having either depression or anxiety.
Tryptophan metabolites, especially serotonin (5-hydroxytryptamine; 5-HT), are considered to be very important for etiology and treatment of depression. Serotonergic neurons in the central nervous system (CNS) are involved in regular behavioral states and physiological processes including arousal, sleep, appetite, pain, release of hormones and mood [8] [9] . Dysfunction of serotonin neurons may lead to depression and other neural disorders [10] [11] [12] . Pharmacological manipulation can successfully increase 5-HT availability in 5-HT neurotransmission in the CNS.
Several biochemical processes intrinsic to the 5-HT neurons were considered to be effectively manipulated by actions of chemical substances including the loading of a precursor amino acid, tryptophan or 5-hydroxytryptophan (5-HTP) inhibiting degradative enzymes, the monoamine oxydases; inducing release of 5-HT; inhibiting 5-HT transport or uptake; blocking autoreceptors at the terminals or cell bodies as well as agonists activating post synaptic receptors.
Recently, however, the serotonin theory of depression has been questioned as reviewed by Moncrief J. et al. [13]
For example, two meta-analyses indicated that 5-HIAA levels in the cerebrospinal fluid showed no association with depression. Plasma levels of serotonin were also shown to have no relationship with depression. Two meta-analyses showed the 5-HT1A receptor and SERT (serotonin reuptake transporter) binding showed weak and inconsistent evidence of reduced binding in some areas. One meta-analysis of tryptophan depletion studies found no effect in most healthy volunteers. The SERT genes such as (5-HTTLPR) revealed no evidence of an association with depression or of an interaction between genotype, stress and depression. The main areas of serotonin research provided no consistent evidence of there being an association between serotonin and depression. Furthermore, it was shown that 5-HT synthesis in suicides in the brainstem is higher than healthy people [14] .
We recently showed that there are no significant differences between plasma levels of TRP between HC and MMD (major monopolar depression). Plasma levels of TRP of HC (healthy controls) are higher in young men, young women, old men, and old women in this order. Serotonin (5-HT) levels are higher in MMD than HC. Plasma levels of 5-HIAA of HC are also higher than those of patients of MMD. Plasma levels of kynurenine (KYN) of healthy old men and old women are higher than those of young men and old women. Plasma levels of KYN are higher in old women and young men of MMD than those of HC [15] [16] [17] .
We also studied about tryptophan metabolites levels in bipolar depression (BD). Plasma levels of TRP are not different between HC and patients of BDII (type II BD). Serotonin (5-HT) levels are higher in BDII than HC. Plasma levels of 5-HIAA of HC are higher than those of old women of BDII, but lower in young women of BDII. Plasma levels of kynurenine (KYN) of HC are not different from those of patients of BDII [18] .
These results suggest that tryptophan metabolites such as serotonin or kynurenine may be produced by gut microbiota and may have nothing to do with biochemical changes in the brain in patients of depression.
2.2. Transport of Amino Acids from the Blood to the Brain
Amino acids are important substances that must be transported to tissues such as the brain and muscles. The process is considered insulin dependent. We want to know whether some important amino acids are transported differently from other amino acids. Especially tryptophan is important because it is converted to serotonin, melatonin or kynurenine. Results showed that Amino acids levels in the plasma were measured after the intakes of 50 grams of glucose or sucrose to young (18 - 22 years old) and old (≥ 50 years old) men. Total amino acids in the plasma decreased after the intakes of glucose. Total and non-essential amino acids in the plasma decreased significantly at 120 min after the intakes of glucose in young and old men, but only sucrose caused their decreases in both aged and young men. Both glucose and sucrose intakes decreased significantly the plasma levels of the total essential and branched amino acids in young and old men. Surprisingly, plasma levels of tryptophan did not decrease upon the administration of glucose but only slightly decreased upon the administration of sucrose in young men. It is shown that not all the amino acids were transported well into tissues upon the administration of glucose or sucrose. Tryptophan seems to be relatively resistant for insulin to facilitate the transportation into tissues [19] [20] .
As to the trans port of tryptophan to the brain, Fernstrome, J.D. and Wurtman R.J. indicated that intakes of tryptophan in foods or injection of insulin increased levels of serotonin and tryptophan in the brain [21] [22] . They indicated that carbohydrate ingestion increased the secretion of insulin which raised plasma levels of tryptophan and lowered the concentrations of the competing amino acids such as branched neutral amino acids in rats [22] . As indicated by them [21] [22] , tryptophan is one of the most important substrates for such transmitters as serotonin and melatonin. Since serotonin is known to decrease depression, it is important to know about transport of tryptophan to the brain and tissues. Tryptophan is transported to the brain competitively with other amino acids [21] [22] . Figure 1 shows the transport of tryptophan and other amino acids to the brain and tissues.
2.3. Transport of Tryptophan or Serotonin in Diseased Brain
The endothelial cells of blood vessels in the brain have the blood brain barrier (BBB) which prevents the movement or infusion of substances toxic or hazardous substances. The brain protects from damages caused by such substances. BBB prevents the entrance of useful or important substances for the activity such
![]()
Figure 1. Transport of tryptophan or serotonin in deseased brain.
as drugs needed for mental health.
Recently, it was shown that the BBB was not functional and substances pass through the endothelial cells to the brain stroma.
Complex tight junctions (TJs) between brain ECs constituted by proteins of the claudin (Cldn) family and occludin (Ocln) block the paracellular pathway [23] . Whereas Cldn5 is also found in nonbarrier endothelium, Cldn3 is predominantly present in brain ECs with specific role in the establishment and maintenance of BBB TJ morphology [24] [25] . ECs rapidly lose their barrier and selective transport properties under pathological conditions in vivo and upon cultivation in vitro, indicating that the healthy brain provides inductive and maintenance signals for the BBB.
Blood-brain barrier and intestinal barrier leak in stress and mood disorders. The blood-brain barrier (BBB) is formed by endothelial cells, pericytes and astrocyte end-feet linking to the capillary wall (Figure 2). The restricted permeability between endothelial cells of the BBB is maintained by substances, such as TJ (tight junction) molecules and JAM (junction adhesion molecules). Depression, stress disorders, anxiety have all been associated with increased levels of circulating pro-inflammatory cytokines, such as IL-6, TNF-α, and IL-1β. This caused breakdown of BBB.
As to tryptophan metabolites, especially tryptophan and serotonin, these molecules are produced by gut microbiota and pass through intestinal epithelia and
![]()
Figure 2. Permeability an its changes in many diseases.
enter the blood. In the brain, under disease conditions, they migrate into the brain.
Serotonin is transported by serotonin reuptake transporter [26] , 5 HT4 receptor [27] , and damaged BBB.
Taking the information into account, Figure 3 is presented.
These data indicate that serotonin migrates into diseased brain so that the levels of serotonin in various mental diseases may be higher than in normal brain.
3. Importance of Tryptophan Metabolites in Health and Diseases
Dietary tryptophan is degraded by enzymes of digestive juice or microbiota.
There are three pathways in tryptophan metabolism (Figure 4).
Not much has been known about substances in indole pathway. Serotonin pathway is by far the most known pathway because serotonin (important for mental health or other physiological functions) and melatonin (important for a circadian rhythm) are involved.
Table 1 shows effects of tryptophan metabolites in diseases.
Many tryptophan metabolites play important roles in health and diseases which will be elucidated in details in the future.
Recently much attention has been paid to kynurenine because kynurenine inhibits T cell functions, thus causes tumor growth.
IDO, which converts tryptophan to kynurenine, was found to be broadly expressed in human tumors [28] [29] [30] and thought to bring about cancer development primarily by tumor immune escape [31] [32] [33] [34] . T cells are very sensitive to low tryptophan levels and cell death under tryptophan deprivation conditions [35] . Low tryptophan and kynurenine metabolites cause effector
![]()
Figure 3. Transport of tryptophan and serotonin from the intestinal cavity to the brain.
![]()
Figure 4. Pathways of tryptophan metabolism
![]()
Table 1. Effects of tryptophan metabolites.
T cell anergy, decrease tumor immune cell infiltration and increase regulatory to effector T cell ratio [36] [37] .
In the field of immunotherapy of cancer, roles of KYN pathway have been studied extensively. Some tumors express high levels of the PD-1 (programmed cell death-1)-biding ligand (PD-L1), and initial trials of anti-PD-1 therapy found that PD-L1 expression correlated well with response to therapy [38] . In order for checkpoint therapy more effective, IDO has been used. The rationale for the use of IDOI is that kynurenine is immunosuppressive, thus preventing effective T-cell attacks to tumors
To discuss more about functions of other metabolites may be beyond the scope of this review.
4. Autism
Autism is very controversial among many psychological disorders. Some people affected by autism are completely disabled, while other people can enjoy high quality of life. Since autism has a wide range of symptoms, it is called autism spectrum disorder, or ASD. As many other psychiatric diseases, the number of people affected by ASD is increasing 10 fold increase in the past 40 years. Generally recognized symptoms of autism are difficulty socializing and repetitive behaviors. However some people ASD show excellent ability in math, music or art. Since some symptoms are not bothering daily lives, people do not recognize that they have symptoms of autism.
As to genetic components of autism, many genes have been identified to be involved in autism, but most of them are weak in detection of autism. Only 15% of genetic autism cases can be directly attributed to changes in genes [39] .
40% to 50% of people with autism suffer from depression and anxiety, which rate is two to four times greater than that of general population [40] . Some 50% - 80% of people with ASD suffer from gut dysbiosis [41] . It has been shown that not only people with autism, but their close relatives have a high level of gastrointestinal symptoms.
Hereditary factors may be related to gut permeability.
Numerous studies have demonstrated the relation between gut microbiota and ASD [40] , I want discuss relationship between tryptophan metabolites, such as tryptophan or serotonin, in ASD.
Serotonin has existed as a signaling molecule across phylogeny [42] . More than 50 years ago whole blood serotonin levels have been shown in a subset of children with autism have been shown. Serotonin has been found to be important for social function, repetitive behavior and sensory development. Genetic linkage studies of whole blood serotonin levels and ASD risk indicate chromosomal region having the serotonin transporter (SERT) gen in males but not in females [43] . A knock-in mouse model of one of SERT genes variants show s increased serotonin clearance, increased serotonin receptor sensitivity, and repetitive behaviors. These results indicate importance of serotonin in many behaviors such as repetitive behavior in ASD.
What roles kynurenine pathways play in the brain of autistic patients?
Relationship between neuroinflammation and kynurenine pathway has attracted attention of researchers because increased frequency of autoimmune disease, allergies, infections have been shown in both autism patients and their parents [44] [45] . The persistence of immune inflammatory deregulation may result in mitochondrial dysfunction and oxidative stress. Chronic inflammation activate the kynurenine pathway which increase in neurotoxic metabolites and cytotoxicity, causing changes in the glutamate system.
As shown in Figure 5 activated gut microbiota may degrades tryptophan leading to generation of kynurenine, further Quinolinic acid (Quin) or kynurenic acid (Kyna). Together with materials in oxidative and nitorosative stress ROS and NOS damages the functions of nerve cells and glia in the brain. The activation of kynurenine pathways may result the activation of glutamic nerves which may cause aberrant behaviors [45] .
![]()
ROS; reactive oxygen species, NOS; nitric acid species.
Figure 5. Kynurenine pathway and effects on mitochondria and brain.
5. Diabetes Mellitus (DM)
There are two types diabetes, type 1 and 2. Compared to type 1 DM, where insulin deficiency is seen, type two shows a combination of insulin deficiency and insulin resistance. Various factors such as sedentary life work, visceral obesity, lack of exercise, poor dietary habits, and genetic factors contribute toward increasing incidences of type2 (T2) DM [46] . Obesity has been shown to increase T2DM by decreasing insulin sensitivity in adipose tissues, liver, skeletal muscle resulting in impaired β cell function [47] .
Data from World Health Organization (WHO) shows that the number of people with diabetes increased from 108 million in 1980 to 422 million in 2014. The global prevalence among adults over 18 years of age increased from 4.7% in 1980 to 8.5% in 2014. It was noticed that between 2000 and 2016 there was a 5% increase in premature mortality from diabetes [48] .
Depression occurs two to three times higher in people with diabetes mellitus, the majority of the cases remaining under-diagnosed. It is important to identify depression in diabetic patients and identify the possible ways to address both diseases. Possible common pathophysiological mechanisms are stress and inflammation, while emphasis was made on screening for depression in diabetic patients [49] .
The microbiota of persons with obesity is highyly efficient in absorbing fats and sugars. This is seen in patients of DM.
We measured tryptophan metabolites of patients of T2DM [50] .
Tryptophan metabolites in plasma samples from 20 male subjects with type 2 diabetes mellitus (T2DM) and 20 nondiabetic reference males were analyzed by ultra high performance liquid chromatography. Tryptophan levels in the diabetic subjects were significantly lower than those in nondiabetic subjects. The concentrations of 5-hydroxytryptophan, 5-hydroxyindoleacetic acid, kynurenic acid, 3-hydroxykynurenine, 3-hydroxyanthranilic acid, and xanthurenic acid were found to be higher in the diabetic patients. These results suggest that tryptophan was metabolized more in T2DM patients than in nondiabetic subjects. In the kynurenine pathway, the degradation of tryptophan seems to be accelerated in patients with higher plasma levels of tryptophan than in patients with lower levels of tryptophan. In the serotonin pathway, when the level of tryptophan is low, the conversion of serotonin to 5-hydroxyindoleacetic acid appears to be accelerated. In conclusion, our results suggest that T2DM patients may be exposed to stress constantly.
6. Heart Disease
Heart disease is closely linked depression. People with heart disease experience depression more than others. People who survive heart attacks feel depressed 6 times more than the general population. To have heart disease and depression increase the mortality rate more than twice.
Patients with affective disorder may have a higher rate of mortality from heart disease compared with normal controls; recent data on heart rate variability abnormalities in depressed patients may be a clue to the mechanism of the increased risk [51] .
Gut microbiota seems to play a role in the health of heart. Methylamine N-oxide (TMAO) is known to cause atherosclerosis. When gut microbiomes digest meat, TMAO is produced. This may be caused increasing dangers of heart attacks
7. Conclusions
Tremendous amounts of research are going on about roles of microbiota in health and diseases. Microbiota seems t be causes of almost all of human diseases.
I am sure that more research will elucidate mechanisms of disease and therapeutic uses of microbiota and their metabolites.